• Campbell Biologie Francais Pdf To Jpg Online

Acknowledgement Of Service Form D10 Pdf Reader Anyone purchased from e. D Body for $1. I can also confirm that the website is legit. My brother ordered a Canon EOS 5. Campbell Biologie Francais Pdf Merge. Fill Purves Biologie Pdf, download blank or editable online. Sign, fax and printable from PC, iPad, tablet or mobile with PDFfiller Instantly No software. Muscle Contraction Steps Skeletal Muscle Anatomy Muscular System Science Biology Medical Science Nursing Process Human Body Systems Nursing School Tips Medical School Flow chart displaying how smooth muscle and skeletal muscle contracts.

The cell (from Latincella, meaning 'small room'^[1]) is the basic structural, functional, and biological unit of all known organisms. A cell is the smallest unit of life. Cells are often called the 'building blocks of life'. The study of cells is called cell biology or cellular biology.

Cells consist of cytoplasm enclosed within a membrane, which contains many biomolecules such as proteins and nucleic acids.^[2] Organisms can be classified as unicellular (consisting of a single cell; including bacteria) or multicellular (including plants and animals).^[3] The number of cells in plants and animals varies from species to species, it has been estimated that humans contain somewhere around 40 trillion (4×10¹³) cells.^[a][4]Most plant and animal cells are visible only under a microscope, with dimensions between 1 and 100 micrometres.^[5]

Cells were discovered by Robert Hooke in 1665, who named them for their resemblance to cells inhabited by Christian monks in a monastery.^[6][7]Cell theory, first developed in 1839 by Matthias Jakob Schleiden and Theodor Schwann, states that all organisms are composed of one or more cells, that cells are the fundamental unit of structure and function in all living organisms, and that all cells come from pre-existing cells.^[8] Cells emerged on Earth at least 3.5 billion years ago.^[9][10][11]

• 1Cell types
• 2Subcellular components
  • 2.4Organelles
• 3Structures outside the cell membrane
  • 3.2Prokaryotic
• 4Cellular processes
• 5Multicellularity
• 6Origins

Cell types

Cells are of two types: eukaryotic, which contain a nucleus, and prokaryotic, which do not. Prokaryotes are single-celled organisms, while eukaryotes can be either single-celled or multicellular.

Prokaryotic cells

Prokaryotes include bacteria and archaea, two of the threedomains of life. Prokaryotic cells were the first form of life on Earth, characterised by having vital biological processes including cell signaling. They are simpler and smaller than eukaryotic cells, and lack membrane-bound organelles such as a nucleus. The DNA of a prokaryotic cell consists of a single circular chromosome that is in direct contact with the cytoplasm. The nuclear region in the cytoplasm is called the nucleoid. Most prokaryotes are the smallest of all organisms ranging from 0.5 to 2.0 µm in diameter.^[12]

A prokaryotic cell has three architectural regions:

• Enclosing the cell is the cell envelope – generally consisting of a plasma membrane covered by a cell wall which, for some bacteria, may be further covered by a third layer called a capsule. Though most prokaryotes have both a cell membrane and a cell wall, there are exceptions such as Mycoplasma (bacteria) and Thermoplasma (archaea) which only possess the cell membrane layer. The envelope gives rigidity to the cell and separates the interior of the cell from its environment, serving as a protective filter. The cell wall consists of peptidoglycan in bacteria, and acts as an additional barrier against exterior forces. It also prevents the cell from expanding and bursting (cytolysis) from osmotic pressure due to a hypotonic environment. Some eukaryotic cells (plant cells and fungal cells) also have a cell wall.
• Inside the cell is the cytoplasmic region that contains the genome (DNA), ribosomes and various sorts of inclusions.^[3] The genetic material is freely found in the cytoplasm. Prokaryotes can carry extrachromosomal DNA elements called plasmids, which are usually circular. Linear bacterial plasmids have been identified in several species of spirochete bacteria, including members of the genus Borrelia notably Borrelia burgdorferi, which causes Lyme disease.^[13] Though not forming a nucleus, the DNA is condensed in a nucleoid. Plasmids encode additional genes, such as antibiotic resistance genes.
• On the outside, flagella and pili project from the cell's surface. These are structures (not present in all prokaryotes) made of proteins that facilitate movement and communication between cells.

Eukaryotic cells

Plants, animals, fungi, slime moulds, protozoa, and algae are all eukaryotic. These cells are about fifteen times wider than a typical prokaryote and can be as much as a thousand times greater in volume. The main distinguishing feature of eukaryotes as compared to prokaryotes is compartmentalization: the presence of membrane-bound organelles (compartments) in which specific activities take place. Most important among these is a cell nucleus,^[3] an organelle that houses the cell's DNA. This nucleus gives the eukaryote its name, which means 'true kernel (nucleus)'. Other differences include:

• The plasma membrane resembles that of prokaryotes in function, with minor differences in the setup. Cell walls may or may not be present.
• The eukaryotic DNA is organized in one or more linear molecules, called chromosomes, which are associated with histone proteins. All chromosomal DNA is stored in the cell nucleus, separated from the cytoplasm by a membrane.^[3] Some eukaryotic organelles such as mitochondria also contain some DNA.
• Many eukaryotic cells are ciliated with primary cilia. Primary cilia play important roles in chemosensation, mechanosensation, and thermosensation. Each cilium may thus be 'viewed as a sensory cellular antennae that coordinates a large number of cellular signaling pathways, sometimes coupling the signaling to ciliary motility or alternatively to cell division and differentiation.'^[14]
• Motile eukaryotes can move using motile cilia or flagella. Motile cells are absent in conifers and flowering plants.^[15] Eukaryotic flagella are more complex than those of prokaryotes.^[16]

Subcellular components

All cells, whether prokaryotic or eukaryotic, have a membrane that envelops the cell, regulates what moves in and out (selectively permeable), and maintains the electric potential of the cell. Inside the membrane, the cytoplasm takes up most of the cell's volume. All cells (except red blood cells which lack a cell nucleus and most organelles to accommodate maximum space for hemoglobin) possess DNA, the hereditary material of genes, and RNA, containing the information necessary to build various proteins such as enzymes, the cell's primary machinery. There are also other kinds of biomolecules in cells. This article lists these primary cellular components, then briefly describes their function.

Membrane

The cell membrane, or plasma membrane, is a biological membrane that surrounds the cytoplasm of a cell. In animals, the plasma membrane is the outer boundary of the cell, while in plants and prokaryotes it is usually covered by a cell wall. This membrane serves to separate and protect a cell from its surrounding environment and is made mostly from a double layer of phospholipids, which are amphiphilic (partly hydrophobic and partly hydrophilic). Hence, the layer is called a phospholipid bilayer, or sometimes a fluid mosaic membrane. Embedded within this membrane is a variety of protein molecules that act as channels and pumps that move different molecules into and out of the cell.^[3] The membrane is semi-permeable, and selectively permeable, in that it can either let a substance (molecule or ion) pass through freely, pass through to a limited extent or not pass through at all. Cell surface membranes also contain receptor proteins that allow cells to detect external signaling molecules such as hormones.

Cytoskeleton

The cytoskeleton acts to organize and maintain the cell's shape; anchors organelles in place; helps during endocytosis, the uptake of external materials by a cell, and cytokinesis, the separation of daughter cells after cell division; and moves parts of the cell in processes of growth and mobility. The eukaryotic cytoskeleton is composed of microfilaments, intermediate filaments and microtubules. There are a great number of proteins associated with them, each controlling a cell's structure by directing, bundling, and aligning filaments.^[3] The prokaryotic cytoskeleton is less well-studied but is involved in the maintenance of cell shape, polarity and cytokinesis.^[18] The subunit protein of microfilaments is a small, monomeric protein called actin. The subunit of microtubules is a dimeric molecule called tubulin. Intermediate filaments are heteropolymers whose subunits vary among the cell types in different tissues. But some of the subunit protein of intermediate filaments include vimentin, desmin, lamin (lamins A, B and C), keratin (multiple acidic and basic keratins), neurofilament proteins (NF–L, NF–M).

Genetic material

Two different kinds of genetic material exist: deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). Cells use DNA for their long-term information storage. The biological information contained in an organism is encoded in its DNA sequence.^[3] RNA is used for information transport (e.g., mRNA) and enzymatic functions (e.g., ribosomal RNA). Transfer RNA (tRNA) molecules are used to add amino acids during protein translation.

Prokaryotic genetic material is organized in a simple circular bacterial chromosome in the nucleoid region of the cytoplasm. Eukaryotic genetic material is divided into different,^[3] linear molecules called chromosomes inside a discrete nucleus, usually with additional genetic material in some organelles like mitochondria and chloroplasts (see endosymbiotic theory).

A human cell has genetic material contained in the cell nucleus (the nuclear genome) and in the mitochondria (the mitochondrial genome). In humans the nuclear genome is divided into 46 linear DNA molecules called chromosomes, including 22 homologous chromosome pairs and a pair of sex chromosomes. The mitochondrial genome is a circular DNA molecule distinct from the nuclear DNA. Although the mitochondrial DNA is very small compared to nuclear chromosomes,^[3] it codes for 13 proteins involved in mitochondrial energy production and specific tRNAs.

Foreign genetic material (most commonly DNA) can also be artificially introduced into the cell by a process called transfection. This can be transient, if the DNA is not inserted into the cell's genome, or stable, if it is. Certain viruses also insert their genetic material into the genome.

Organelles

Organelles are parts of the cell which are adapted and/or specialized for carrying out one or more vital functions, analogous to the organs of the human body (such as the heart, lung, and kidney, with each organ performing a different function).^[3] Both eukaryotic and prokaryotic cells have organelles, but prokaryotic organelles are generally simpler and are not membrane-bound.

There are several types of organelles in a cell. Some (such as the nucleus and golgi apparatus) are typically solitary, while others (such as mitochondria, chloroplasts, peroxisomes and lysosomes) can be numerous (hundreds to thousands). The cytosol is the gelatinous fluid that fills the cell and surrounds the organelles.

Eukaryotic

• Cell nucleus: A cell's information center, the cell nucleus is the most conspicuous organelle found in a eukaryotic cell. It houses the cell's chromosomes, and is the place where almost all DNA replication and RNA synthesis (transcription) occur. The nucleus is spherical and separated from the cytoplasm by a double membrane called the nuclear envelope. The nuclear envelope isolates and protects a cell's DNA from various molecules that could accidentally damage its structure or interfere with its processing. During processing, DNA is transcribed, or copied into a special RNA, called messenger RNA (mRNA). This mRNA is then transported out of the nucleus, where it is translated into a specific protein molecule. The nucleolus is a specialized region within the nucleus where ribosome subunits are assembled. In prokaryotes, DNA processing takes place in the cytoplasm.^[3]
• Mitochondria and Chloroplasts: generate energy for the cell. Mitochondria are self-replicating organelles that occur in various numbers, shapes, and sizes in the cytoplasm of all eukaryotic cells.^[3]Respiration occurs in the cell mitochondria, which generate the cell's energy by oxidative phosphorylation, using oxygen to release energy stored in cellular nutrients (typically pertaining to glucose) to generate ATP. Mitochondria multiply by binary fission, like prokaryotes. Chloroplasts can only be found in plants and algae, and they capture the sun's energy to make carbohydrates through photosynthesis.

• Endoplasmic reticulum: The endoplasmic reticulum (ER) is a transport network for molecules targeted for certain modifications and specific destinations, as compared to molecules that float freely in the cytoplasm. The ER has two forms: the rough ER, which has ribosomes on its surface that secrete proteins into the ER, and the smooth ER, which lacks ribosomes.^[3] The smooth ER plays a role in calcium sequestration and release.
• Golgi apparatus: The primary function of the Golgi apparatus is to process and package the macromolecules such as proteins and lipids that are synthesized by the cell.
• Lysosomes and Peroxisomes: Lysosomes contain digestive enzymes (acid hydrolases). They digest excess or worn-out organelles, food particles, and engulfed viruses or bacteria. Peroxisomes have enzymes that rid the cell of toxic peroxides. The cell could not house these destructive enzymes if they were not contained in a membrane-bound system.^[3]
• Centrosome: the cytoskeleton organiser: The centrosome produces the microtubules of a cell – a key component of the cytoskeleton. It directs the transport through the ER and the Golgi apparatus. Centrosomes are composed of two centrioles, which separate during cell division and help in the formation of the mitotic spindle. A single centrosome is present in the animal cells. They are also found in some fungi and algae cells.
• Vacuoles: Vacuoles sequester waste products and in plant cells store water. They are often described as liquid filled space and are surrounded by a membrane. Some cells, most notably Amoeba, have contractile vacuoles, which can pump water out of the cell if there is too much water. The vacuoles of plant cells and fungal cells are usually larger than those of animal cells.

Eukaryotic and prokaryotic

• Ribosomes: The ribosome is a large complex of RNA and protein molecules.^[3] They each consist of two subunits, and act as an assembly line where RNA from the nucleus is used to synthesise proteins from amino acids. Ribosomes can be found either floating freely or bound to a membrane (the rough endoplasmatic reticulum in eukaryotes, or the cell membrane in prokaryotes).^[19]

Structures outside the cell membrane

Many cells also have structures which exist wholly or partially outside the cell membrane. These structures are notable because they are not protected from the external environment by the semipermeable cell membrane. In order to assemble these structures, their components must be carried across the cell membrane by export processes.

Cell wall

Many types of prokaryotic and eukaryotic cells have a cell wall. The cell wall acts to protect the cell mechanically and chemically from its environment, and is an additional layer of protection to the cell membrane. Different types of cell have cell walls made up of different materials; plant cell walls are primarily made up of cellulose, fungi cell walls are made up of chitin and bacteria cell walls are made up of peptidoglycan.

Campbell Biologie Francais Pdf To Jpg Online

Prokaryotic

Capsule

A gelatinous capsule is present in some bacteria outside the cell membrane and cell wall. The capsule may be polysaccharide as in pneumococci, meningococci or polypeptide as Bacillus anthracis or hyaluronic acid as in streptococci.Capsules are not marked by normal staining protocols and can be detected by India ink or methyl blue; which allows for higher contrast between the cells for observation.^[20]:87

Flagella

Flagella are organelles for cellular mobility. The bacterial flagellum stretches from cytoplasm through the cell membrane(s) and extrudes through the cell wall. They are long and thick thread-like appendages, protein in nature. A different type of flagellum is found in archaea and a different type is found in eukaryotes.

Fimbria

A fimbria also known as a pilus is a short, thin, hair-like filament found on the surface of bacteria. Fimbriae, or pili are formed of a protein called pilin (antigenic) and are responsible for attachment of bacteria to specific receptors of human cell (cell adhesion). There are special types of specific pili involved in bacterial conjugation.

Cellular processes

Replication

Cell division involves a single cell (called a mother cell) dividing into two daughter cells. This leads to growth in multicellular organisms (the growth of tissue) and to procreation (vegetative reproduction) in unicellular organisms. Prokaryotic cells divide by binary fission, while eukaryotic cells usually undergo a process of nuclear division, called mitosis, followed by division of the cell, called cytokinesis. A diploid cell may also undergo meiosis to produce haploid cells, usually four. Haploid cells serve as gametes in multicellular organisms, fusing to form new diploid cells.

DNA replication, or the process of duplicating a cell's genome,^[3] always happens when a cell divides through mitosis or binary fission. This occurs during the S phase of the cell cycle.

In meiosis, the DNA is replicated only once, while the cell divides twice. DNA replication only occurs before meiosis I. DNA replication does not occur when the cells divide the second time, in meiosis II.^[21] Replication, like all cellular activities, requires specialized proteins for carrying out the job.^[3]

Growth and metabolism

Between successive cell divisions, cells grow through the functioning of cellular metabolism. Cell metabolism is the process by which individual cells process nutrient molecules. Metabolism has two distinct divisions: catabolism, in which the cell breaks down complex molecules to produce energy and reducing power, and anabolism, in which the cell uses energy and reducing power to construct complex molecules and perform other biological functions.Complex sugars consumed by the organism can be broken down into simpler sugar molecules called monosaccharides such as glucose. Once inside the cell, glucose is broken down to make adenosine triphosphate (ATP),^[3] a molecule that possesses readily available energy, through two different pathways.

Protein synthesis

Cells are capable of synthesizing new proteins, which are essential for the modulation and maintenance of cellular activities. This process involves the formation of new protein molecules from amino acid building blocks based on information encoded in DNA/RNA. Protein synthesis generally consists of two major steps: transcription and translation.

Transcription is the process where genetic information in DNA is used to produce a complementary RNA strand. This RNA strand is then processed to give messenger RNA (mRNA), which is free to migrate through the cell. mRNA molecules bind to protein-RNA complexes called ribosomes located in the cytosol, where they are translated into polypeptide sequences. The ribosome mediates the formation of a polypeptide sequence based on the mRNA sequence. The mRNA sequence directly relates to the polypeptide sequence by binding to transfer RNA (tRNA) adapter molecules in binding pockets within the ribosome. The new polypeptide then folds into a functional three-dimensional protein molecule.

Motility

Unicellular organisms can move in order to find food or escape predators. Common mechanisms of motion include flagella and cilia.

In multicellular organisms, cells can move during processes such as wound healing, the immune response and cancer metastasis. For example, in wound healing in animals, white blood cells move to the wound site to kill the microorganisms that cause infection. Cell motility involves many receptors, crosslinking, bundling, binding, adhesion, motor and other proteins.^[22] The process is divided into three steps – protrusion of the leading edge of the cell, adhesion of the leading edge and de-adhesion at the cell body and rear, and cytoskeletal contraction to pull the cell forward. Each step is driven by physical forces generated by unique segments of the cytoskeleton.^[23][24]

Multicellularity

Cell specialization

Multicellular organisms are organisms that consist of more than one cell, in contrast to single-celled organisms.^[25]

In complex multicellular organisms, cells specialize into different cell types that are adapted to particular functions. In mammals, major cell types include skin cells, muscle cells, neurons, blood cells, fibroblasts, stem cells, and others. Cell types differ both in appearance and function, yet are genetically identical. Cells are able to be of the same genotype but of different cell type due to the differential expression of the genes they contain.

Most distinct cell types arise from a single totipotent cell, called a zygote, that differentiates into hundreds of different cell types during the course of development. Differentiation of cells is driven by different environmental cues (such as cell–cell interaction) and intrinsic differences (such as those caused by the uneven distribution of molecules during division).

Origin of multicellularity

Multicellularity has evolved independently at least 25 times,^[26] including in some prokaryotes, like cyanobacteria, myxobacteria, actinomycetes, Magnetoglobus multicellularis or Methanosarcina. However, complex multicellular organisms evolved only in six eukaryotic groups: animals, fungi, brown algae, red algae, green algae, and plants.^[27] It evolved repeatedly for plants (Chloroplastida), once or twice for animals, once for brown algae, and perhaps several times for fungi, slime molds, and red algae.^[28] Multicellularity may have evolved from colonies of interdependent organisms, from cellularization, or from organisms in symbiotic relationships.

The first evidence of multicellularity is from cyanobacteria-like organisms that lived between 3 and 3.5 billion years ago.^[26] Other early fossils of multicellular organisms include the contested Grypania spiralis and the fossils of the black shales of the PalaeoproterozoicFrancevillian Group Fossil B Formation in Gabon.^[29]

The evolution of multicellularity from unicellular ancestors has been replicated in the laboratory, in evolution experiments using predation as the selective pressure.^[26]

Origins

The origin of cells has to do with the origin of life, which began the history of life on Earth.

Origin of the first cell

There are several theories about the origin of small molecules that led to life on the early Earth. They may have been carried to Earth on meteorites (see Murchison meteorite), created at deep-sea vents, or synthesized by lightning in a reducing atmosphere (see Miller–Urey experiment). There is little experimental data defining what the first self-replicating forms were. RNA is thought to be the earliest self-replicating molecule, as it is capable of both storing genetic information and catalyzing chemical reactions (see RNA world hypothesis), but some other entity with the potential to self-replicate could have preceded RNA, such as clay or peptide nucleic acid.^[30]

Cells emerged at least 3.5 billion years ago.^[9][10][11] The current belief is that these cells were heterotrophs. The early cell membranes were probably more simple and permeable than modern ones, with only a single fatty acid chain per lipid. Lipids are known to spontaneously form bilayered vesicles in water, and could have preceded RNA, but the first cell membranes could also have been produced by catalytic RNA, or even have required structural proteins before they could form.^[31]

Origin of eukaryotic cells

The eukaryotic cell seems to have evolved from a symbiotic community of prokaryotic cells. DNA-bearing organelles like the mitochondria and the chloroplasts are descended from ancient symbiotic oxygen-breathing proteobacteria and cyanobacteria, respectively, which were endosymbiosed by an ancestral archaean prokaryote.

There is still considerable debate about whether organelles like the hydrogenosome predated the origin of mitochondria, or vice versa: see the hydrogen hypothesis for the origin of eukaryotic cells.

History of research

• 1632–1723: Antonie van Leeuwenhoek taught himself to make lenses, constructed basic optical microscopes and drew protozoa, such as Vorticella from rain water, and bacteria from his own mouth.
• 1665: Robert Hooke discovered cells in cork, then in living plant tissue using an early compound microscope. He coined the term cell (from Latincella, meaning 'small room'^[1]) in his book Micrographia (1665).^[32]
• 1839: Theodor Schwann and Matthias Jakob Schleiden elucidated the principle that plants and animals are made of cells, concluding that cells are a common unit of structure and development, and thus founding the cell theory.
• 1855: Rudolf Virchow stated that new cells come from pre-existing cells by cell division (omnis cellula ex cellula).
• 1859: The belief that life forms can occur spontaneously (generatio spontanea) was contradicted by Louis Pasteur (1822–1895) (although Francesco Redi had performed an experiment in 1668 that suggested the same conclusion).
• 1931: Ernst Ruska built the first transmission electron microscope (TEM) at the University of Berlin. By 1935, he had built an EM with twice the resolution of a light microscope, revealing previously unresolvable organelles.
• 1953: Based on Rosalind Franklin's work, Watson and Crick made their first announcement on the double helix structure of DNA.
• 1981: Lynn Margulis published Symbiosis in Cell Evolution detailing the endosymbiotic theory.

See also

References

1. ^ ^ab'Cell'. Online Etymology Dictionary. Retrieved 31 December 2012.
2. ^Cell Movements and the Shaping of the Vertebrate Body in Chapter 21 of Molecular Biology of the Cell fourth edition, edited by Bruce Alberts (2002) published by Garland Science.
   The Alberts text discusses how the 'cellular building blocks' move to shape developing embryos. It is also common to describe small molecules such as amino acids as 'molecular building blocks'.
3. ^ ^{abcdefghijklmnopqr} This article incorporates public domain material from the NCBI document 'What Is a Cell?'. 30 March 2004.
4. ^ ^abcBianconi, Eva; Piovesan, Allison; Facchin, Federica; Beraudi, Alina; Casadei, Raffaella; Frabetti, Flavia; Vitale, Lorenza; Pelleri, Maria Chiara; Tassani, Simone (November 2013). 'An estimation of the number of cells in the human body'. Annals of Human Biology. 40 (6): 463–471. doi:10.3109/03014460.2013.807878. ISSN0301-4460. These partial data correspond to a total number of 3.72±0.81×10¹³ [cells].
5. ^Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life. Boston, Massachusetts: Pearson Prentice Hall. ISBN9780132508827.
6. ^Karp, Gerald (19 October 2009). Cell and Molecular Biology: Concepts and Experiments. John Wiley & Sons. p. 2. ISBN9780470483374. Hooke called the pores cells because they reminded him of the cells inhabited by monks living in a monastery.
7. ^Tero AC (1990). Achiever's Biology. Allied Publishers. p. 36. ISBN9788184243697. In 1665, an Englishman, Robert Hooke observed a thin slice of' cork under a simple microscope. (A simple microscope is a microscope with only one biconvex lens, rather like a magnifying glass). He saw many small box like structures. These reminded him of small rooms called 'cells' in which Christian monks lived and meditated.
8. ^Maton A (1997). Cells Building Blocks of Life. New Jersey: Prentice Hall. ISBN9780134234762.
9. ^ ^abSchopf JW, Kudryavtsev AB, Czaja AD, Tripathi AB (2007). 'Evidence of Archean life: Stromatolites and microfossils'. Precambrian Research. 158 (3–4): 141–55. Bibcode:2007PreR.158.141S. doi:10.1016/j.precamres.2007.04.009.
10. ^ ^abSchopf JW (2006). 'Fossil evidence of Archaean life'. Philos Trans R Soc Lond B Biol Sci. 29 (361(1470)): 869–885. doi:10.1098/rstb.2006.1834. PMC1578735. PMID16754604.
11. ^ ^abRaven PH, Johnson GB (2002). Biology. McGraw-Hill Education. p. 68. ISBN9780071122610. Retrieved 7 July 2013.
12. ^Microbiology : Principles and Explorations By Jacquelyn G. Black
13. ^European Bioinformatics Institute, Karyn's Genomes: Borrelia burgdorferi, part of 2can on the EBI-EMBL database. Retrieved 5 August 2012
14. ^Satir P, Christensen ST (June 2008). 'Structure and function of mammalian cilia'. Histochemistry and Cell Biology. 129 (6): 687–93. doi:10.1007/s00418-008-0416-9. PMC2386530. PMID18365235. 1432-119X.
15. ^PH Raven, Evert RF, Eichhorm SE (1999) Biology of Plants, 6th edition. WH Freeman, New York
16. ^Blair, David F.; Dutcher, Susan K. (1992-01-01). 'Flagella in prokaryotes and lower eukaryotes'. Current Opinion in Genetics & Development. 2 (5): 756–767. doi:10.1016/S0959-437X(05)80136-4. ISSN0959-437X.
17. ^ ^abCampbell Biology—Concepts and Connections. Pearson Education. 2009. p. 320.
18. ^Michie KA, Löwe J (2006). 'Dynamic filaments of the bacterial cytoskeleton'. Annual Review of Biochemistry. 75: 467–92. doi:10.1146/annurev.biochem.75.103004.142452. PMID16756499.
19. ^Ménétret JF, Schaletzky J, Clemons WM, Osborne AR, Skånland SS, Denison C, Gygi SP, Kirkpatrick DS, Park E, Ludtke SJ, Rapoport TA, Akey CW (December 2007). 'Ribosome binding of a single copy of the SecY complex: implications for protein translocation'. Molecular Cell. 28 (6): 1083–92. doi:10.1016/j.molcel.2007.10.034. PMID18158904.
20. ^Prokaryotes. Newnes. Apr 11, 1996. ISBN9780080984735.
21. ^Campbell Biology—Concepts and Connections. Pearson Education. 2009. p. 138.
22. ^Ananthakrishnan R, Ehrlicher A. 'The Forces Behind Cell Movement'. Biolsci.org. Retrieved 2009-04-17.
23. ^Alberts B, Johnson A, Lewis J. et al. Molecular Biology of the Cell, 4e. Garland Science. 2002
24. ^Ananthakrishnan R, Ehrlicher A (June 2007). 'The forces behind cell movement'. International Journal of Biological Sciences. 3 (5): 303–17. doi:10.7150/ijbs.3.303. PMC1893118. PMID17589565.
25. ^Becker WM, et al. (2009). The world of the cell. Pearson Benjamin Cummings. p. 480. ISBN9780321554185.
26. ^ ^abcGrosberg RK, Strathmann RR (2007). 'The evolution of multicellularity: A minor major transition?'(PDF). Annu Rev Ecol Evol Syst. 38: 621–54. doi:10.1146/annurev.ecolsys.36.102403.114735.
27. ^Popper ZA, Michel G, Hervé C, Domozych DS, Willats WG, Tuohy MG, Kloareg B, Stengel DB (2011). 'Evolution and diversity of plant cell walls: from algae to flowering plants'(PDF). Annual Review of Plant Biology. 62: 567–90. doi:10.1146/annurev-arplant-042110-103809. hdl:10379/6762. PMID21351878.
28. ^Bonner JT (1998). 'The Origins of Multicellularity'(PDF). Integrative Biology: Issues, News, and Reviews. 1 (1): 27–36. doi:10.1002/(SICI)1520-6602(1998)1:1<27::AID-INBI4>3.0.CO;2-6. ISSN1093-4391. Archived from the original(PDF, 0.2 MB) on March 8, 2012.Cite uses deprecated parameter deadurl= (help)
29. ^El Albani A, Bengtson S, Canfield DE, Bekker A, Macchiarelli R, Mazurier A, Hammarlund EU, Boulvais P, Dupuy JJ, Fontaine C, Fürsich FT, Gauthier-Lafaye F, Janvier P, Javaux E, Ossa FO, Pierson-Wickmann AC, Riboulleau A, Sardini P, Vachard D, Whitehouse M, Meunier A (July 2010). 'Large colonial organisms with coordinated growth in oxygenated environments 2.1 Gyr ago'. Nature. 466 (7302): 100–04. Bibcode:2010Natur.466.100A. doi:10.1038/nature09166. PMID20596019.
30. ^Orgel LE (December 1998). 'The origin of life – a review of facts and speculations'. Trends in Biochemical Sciences. 23 (12): 491–95. doi:10.1016/S0968-0004(98)01300-0. PMID9868373.
31. ^Griffiths G (December 2007). 'Cell evolution and the problem of membrane topology'. Nature Reviews. Molecular Cell Biology. 8 (12): 1018–24. doi:10.1038/nrm2287. PMID17971839.
32. ^Hooke R (1665). Micrographia: …. London, England: Royal Society of London. p. 113.' … I could exceedingly plainly perceive it to be all perforated and porous, much like a Honey-comb, but that the pores of it were not regular […] these pores, or cells, […] were indeed the first microscopical pores I ever saw, and perhaps, that were ever seen, for I had not met with any Writer or Person, that had made any mention of them before this … ' – Hooke describing his observations on a thin slice of cork. See also: Robert Hooke

Notes

1. ^An approximation made for someone who is 30 years old, weighs 70 kilograms (150 lb), and is 172 centimetres (5.64 ft) tall.^[4] The approximation is not exact, this study estimated that the number of cells was 3.72±0.81×10¹³.^[4]

Further reading

• Alberts B, Johnson A, Lewis J, Morgan D, Raff M, Roberts K, Walter P (2015). Molecular Biology of the Cell (6th ed.). Garland Science. p. 2. ISBN9780815344322.
• Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2014). Molecular Biology of the Cell (6th ed.). Garland. ISBN9780815344322.; The fourth edition is freely available from National Center for Biotechnology Information Bookshelf.
• Lodish H, Berk A, Matsudaira P, Kaiser CA, Krieger M, Scott MP, Zipurksy SL, Darnell J (2004). Molecular Cell Biology (5th ed.). WH Freeman: New York, NY. ISBN9780716743668.
• Cooper GM (2000). The cell: a molecular approach (2nd ed.). Washington, D.C: ASM Press. ISBN9780878931026.

External links

• Inside the Cell – a science education booklet by National Institutes of Health, in PDF and ePub.
• Cell Biology in 'The Biology Project' of University of Arizona.
• The Image & Video Library of The American Society for Cell Biology, a collection of peer-reviewed still images, video clips and digital books that illustrate the structure, function and biology of the cell.
• HighMag Blog, still images of cells from recent research articles.
• New Microscope Produces Dazzling 3D Movies of Live Cells, March 4, 2011 – Howard Hughes Medical Institute.
• WormWeb.org: Interactive Visualization of the C. elegans Cell lineage – Visualize the entire cell lineage tree of the nematode C. elegans

The green fluorescent protein (GFP) is a protein composed of 238 amino acid residues (26.9 kDa) that exhibits bright green fluorescence when exposed to light in the blue to ultraviolet range.^[2][3] Although many other marine organisms have similar green fluorescent proteins, GFP traditionally refers to the protein first isolated from the jellyfishAequorea victoria, avGFP. The GFP from A. victoria has a major excitation peak at a wavelength of 395 nm and a minor one at 475 nm. Its emission peak is at 509 nm, which is in the lower green portion of the visible spectrum. The fluorescence quantum yield (QY) of GFP is 0.79. The GFP from the sea pansy (Renilla reniformis) has a single major excitation peak at 498 nm. GFP makes for an excellent tool in many forms of biology due to its ability to form internal chromophore without requiring any accessory cofactors, gene products, or enzymes / substrates other than molecular oxygen.^[4]

In cell and molecular biology, the GFP gene is frequently used as a reporter of expression.^[5] It has been used in modified forms to make biosensors, and many animals have been created that express GFP, which demonstrates a proof of concept that a gene can be expressed throughout a given organism, in selected organs, or in cells of interest. GFP can be introduced into animals or other species through transgenic techniques, and maintained in their genome and that of their offspring. To date, GFP has been expressed in many species, including bacteria, yeasts, fungi, fish and mammals, including in human cells. Scientists Roger Y. Tsien, Osamu Shimomura, and Martin Chalfie were awarded the 2008 Nobel Prize in Chemistry on 10 October 2008 for their discovery and development of the green fluorescent protein.

• 1History
• 5Applications
  • 5.1Reporter assays

History[edit]

Wild-type GFP (wtGFP)[edit]

In the 1960s and 1970s, GFP, along with the separate luminescent protein aequorin (an enzyme that catalyzes the breakdown of luciferin, releasing light), was first purified from Aequorea victoria and its properties studied by Osamu Shimomura.^[6] In A. victoria, GFP fluorescence occurs when aequorin interacts with Ca²⁺ ions, inducing a blue glow. Some of this luminescent energy is transferred to the GFP, shifting the overall color towards green.^[7] However, its utility as a tool for molecular biologists did not begin to be realized until 1992 when Douglas Prasher reported the cloning and nucleotide sequence of wtGFP in Gene.^[8] The funding for this project had run out, so Prasher sent cDNA samples to several labs. The lab of Martin Chalfie expressed the coding sequence of wtGFP, with the first few amino acids deleted, in heterologous cells of E. coli and C. elegans, publishing the results in Science in 1994.^[9] Frederick Tsuji's lab independently reported the expression of the recombinant protein one month later.^[10] Remarkably, the GFP molecule folded and was fluorescent at room temperature, without the need for exogenous cofactors specific to the jellyfish. Although this near-wtGFP was fluorescent, it had several drawbacks, including dual peaked excitation spectra, pH sensitivity, chloride sensitivity, poor fluorescence quantum yield, poor photostability and poor folding at 37 °C.

The first reported crystal structure of a GFP was that of the S65T mutant by the Remington group in Science in 1996.^[11] One month later, the Phillips group independently reported the wild-type GFP structure in Nature Biotechnology.^[12] These crystal structures provided vital background on chromophore formation and neighboring residue interactions. Researchers have modified these residues by directed and random mutagenesis to produce the wide variety of GFP derivatives in use today. Further research into GFP has shown that it is resistant to detergents, proteases, guanidinium chloride (GdmCl) treatments, and drastic temperature changes.^[13]

GFP derivatives[edit]

Due to the potential for widespread usage and the evolving needs of researchers, many different mutants of GFP have been engineered.^[14][15] The first major improvement was a single point mutation (S65T) reported in 1995 in Nature by Roger Tsien.^[16] This mutation dramatically improved the spectral characteristics of GFP, resulting in increased fluorescence, photostability, and a shift of the major excitation peak to 488 nm, with the peak emission kept at 509 nm. This matched the spectral characteristics of commonly available FITC filter sets, increasing the practicality of use by the general researcher. A 37 °C folding efficiency (F64L) point mutant to this scaffold, yielding enhanced GFP (EGFP), was discovered in 1995 by the laboratories of Thastrup^[17] and Falkow.^[18] EGFP allowed the practical use of GFPs in mammalian cells. EGFP has an extinction coefficient (denoted ε) of 55,000 M⁻¹cm⁻¹.^[19] The fluorescence quantum yield (QY) of EGFP is 0.60. The relative brightness, expressed as ε•QY, is 33,000 M⁻¹cm⁻¹.

Superfolder GFP (sfGFP), a series of mutations that allow GFP to rapidly fold and mature even when fused to poorly folding peptides, was reported in 2006.^[20]

Many other mutations have been made, including color mutants; in particular, blue fluorescent protein (EBFP, EBFP2, Azurite, mKalama1), cyan fluorescent protein (ECFP, Cerulean, CyPet, mTurquoise2), and yellow fluorescent protein derivatives (YFP, Citrine, Venus, YPet). BFP derivatives (except mKalama1) contain the Y66H substitution.They exhibit a broad absorption band in the ultraviolet centered close to 380 nanometers and an emission maximum at 448 nanometers. A green fluorescent protein mutant (BFPms1) that preferentially binds Zn(II) and Cu(II) has been developed. BFPms1 have several important mutations including and the BFP chromophore (Y66H),Y145F for higher quantum yield, H148G for creating a hole into the beta-barrel and several other mutations that increase solubility. Zn(II) binding increases fluorescence intensity, while Cu(II) binding quenches fluorescence and shifts the absorbance maximum from 379 to 444 nm. Therefore, they can be used as Zn biosensor.^[21]

Chromophore binding. The critical mutation in cyan derivatives is the Y66W substitution, which causes the chromophore to form with an indole rather than phenol component. Several additional compensatory mutations in the surrounding barrel are required to restore brightness to this modified chromophore due to the increased bulk of the indole group. In ECFP and Cerulean, the N-terminal half of the seventh strand exhibits two conformations. These conformations both have a complex set of van der Waals interactions with the chromophore. The Y145A and H148D mutations in Cerulean stabilize these interactions and allow the chromophore to be more planar, better packed, and less prone to collisional quenching.^[22]

Additional site-directed random mutagenesis in combination with fluorescence lifetime based screening has further stabilized the seventh β-strand resulting in a bright variant, mTurquoise2, with a quantum yield (QY) of 0.93.^[23] The red-shifted wavelength of the YFP derivatives is accomplished by the T203Y mutation and is due to π-electron stacking interactions between the substituted tyrosine residue and the chromophore.^[3] These two classes of spectral variants are often employed for Förster resonance energy transfer (FRET) experiments. Genetically encoded FRET reporters sensitive to cell signaling molecules, such as calcium or glutamate, protein phosphorylation state, protein complementation, receptor dimerization, and other processes provide highly specific optical readouts of cell activity in real time.

Semirational mutagenesis of a number of residues led to pH-sensitive mutants known as pHluorins, and later super-ecliptic pHluorins. By exploiting the rapid change in pH upon synaptic vesicle fusion, pHluorins tagged to synaptobrevin have been used to visualize synaptic activity in neurons.^[24]

Redox sensitive GFP (roGFP) was engineered by introduction of cysteines into the beta barrel structure. The redox state of the cysteines determines the fluorescent properties of roGFP.^[25]

Nomenclature[edit]

The nomenclature of modified GFPs is often confusing due to overlapping mapping of several GFP versions onto a single name. For example, mGFP often refers to a GFP with an N-terminal palmitoylation that causes the GFP to bind to cell membranes. However, the same term is also used to refer to monomeric GFP, which is often achieved by the dimer interface breaking A206K mutation.^[26] Wild-type GFP has a weak dimerization tendency at concentrations above 5 mg/mL. mGFP also stands for 'modified GFP,' which has been optimized through amino acid exchange for stable expression in plant cells.

In nature[edit]

The purpose of both the (primary) bioluminescence (from aequorin's action on luciferin) and the (secondary) fluorescence of GFP in jellyfish is unknown. GFP is co-expressed with aequorin in small granules around the rim of the jellyfish bell. The secondary excitation peak (480 nm) of GFP does absorb some of the blue emission of aequorin, giving the bioluminescence a more green hue. The serine 65 residue of the GFP chromophore is responsible for the dual-peaked excitation spectra of wild-type GFP. It is conserved in all three GFP isoforms originally cloned by Prasher. Nearly all mutations of this residue consolidate the excitation spectra to a single peak at either 395 nm or 480 nm. The precise mechanism of this sensitivity is complex, but, it seems, involves donation of a hydrogen from serine 65 to glutamate 222, which influences chromophore ionization.^[3] Since a single mutation can dramatically enhance the 480 nm excitation peak, making GFP a much more efficient partner of aequorin, A. victoria appears to evolutionarily prefer the less-efficient, dual-peaked excitation spectrum. Roger Tsien has speculated that varying hydrostatic pressure with depth may affect serine 65's ability to donate a hydrogen to the chromophore and shift the ratio of the two excitation peaks. Thus, the jellyfish may change the color of its bioluminescence with depth. However, a collapse in the population of jellyfish in Friday Harbor, where GFP was originally discovered, has hampered further study of the role of GFP in the jellyfish's natural environment.

Other fluorescent proteins[edit]

There are many GFP-like proteins that, despite being in the same protein family as GFP, are not directly derived from Aequorea victoria. These include dsRed, eqFP611, Dronpa, TagRFPs, KFP, EosFP/IrisFP, Dendra, and so on. Having been developed from proteins in different organisms, these proteins can sometimes display unantipated approaches to chromophore formation. Some of these, such as KFP, are developed from naturally non- or weakly-fluorescent proteins to be greatly improved upon by mutagenesis.^[27] When GFP-like barrels of different spectra characteristics are used, the excitation spectra of one chromophore can be used to power another chromophore (FRET), allowing for conversion between wavelengths of light.^[28]

FMN-binding fluorescent proteins (FbFPs) were developed in 2007 and are a class of small (11-16 kDa), oxygen-independent fluorescent proteins that are derived from blue-light receptors. They are intended especially for the use under anaerobic or hypoxic conditions, since the formation and binding of the Flavin chromophore does not require molecular oxygen, as it is the case with the synthesis of the GFP chromophore.^[29]

Fluorescent proteins with other chromophores, such as UnaG with bilirubin, can display unique properties like red-shifted emission above 600 nm or photoconversion from a green-emitting state to a red-emitting state. They can have excitation and emission wavelengths far enough apart to achieve conversion between red and green light.

A new class of fluorescent protein was evolved from a cyanobacterial (Trichodesmium erythraeum) phycobiliprotein, α-allophycocyanin, and named small ultra red fluorescent protein (smURFP) in 2016. smURFPautocatalytically self-incorporates the chromophorebiliverdin without the need of an external protein, known as a lyase.^[30]Jellyfish- and coral-derived GFP-like proteins require oxygen and produce a stoichiometric amount of hydrogen peroxide upon chromophore formation.^[31]smURFP does not require oxygen or produce hydrogen peroxide and uses the chromophore, bliverdin. smURFP has a large extinction coefficient (180,000 M⁻¹ cm⁻¹) and has a modest quantum yield (0.20), which makes it comparable biophysical brightness to eGFP and ~2-fold brighter than most red or far-red fluorescent proteins derived from coral. smURFP spectral properties are similar to the organic dye Cy5.^[30]

A review of new classes of fluorescent proteins and applications can be found in Trends in Biochemical Sciences.^[32]

Structure[edit]

GFP has a beta barrel structure consisting of eleven β-strands with a pleated sheet arrangement, with an alpha helix containing the covalently bonded chromophore 4-(p-hydroxybenzylidene)imidazolidin-5-one (HBI) running through the center.^[3][11][12] Five shorter alpha helices form caps on the ends of the structure. The beta barrel structure is a nearly perfect cylinder, 42Å long and 24Å in diameter (some studies have reported a diameter of 30Å^[13]),^[11] creating what is referred to as a 'β-can' formation, which is unique to the GFP-like family.^[12] HBI, the spontaneously modified form of the tripeptide Ser65–Tyr66–Gly67, is nonfluorescent in the absence of the properly folded GFP scaffold and exists mainly in the un-ionized phenol form in wtGFP.^[33] Inward-facing sidechains of the barrel induce specific cyclization reactions in Ser65–Tyr66–Gly67 that induce ionization of HBI to the phenolate form and chromophore formation. This process of post-translational modification is referred to as maturation.^[34] The hydrogen-bonding network and electron-stacking interactions with these sidechains influence the color, intensity and photostability of GFP and its numerous derivatives.^[35] The tightly packed nature of the barrel excludes solvent molecules, protecting the chromophore fluorescence from quenching by water. In addition to the auto-cyclization of the Ser65-Tyr66-Gly67, a 1,2-dehydrogenation reaction occurs at the Tyr66 residue.^[13] Besides the three residues that form the chromophore, residues such as Gln94, Arg96, His148, Thr203, and Glu222 all act as stabilizers. The residues of Gln94, Arg96, and His148 are able to stabilize by delocalizing the chromophore charge. Arg96 is the most important stabilizing residue due to the fact that it prompts the necessary structural realignments that are necessary from the HBI ring to occur. Any mutation to the Arg96 residue would result in a decrease in the development rate of the chromophore because proper electrostatic and steric interactions would be lost. Tyr66 is the recipient of hydrogen bonds and does not ionize in order to produce favorable electrostatics.^[36]

Applications[edit]

Reporter assays[edit]

Green fluorescent protein may be used as a reporter gene.^[37][38]

For example, GFP can be used as a reporter for environmental toxicity levels. This protein has been shown to be an effective way to measure the toxicity levels of various chemicals including ethanol, p-formaldehyde, phenol, triclosan, and paraben. GFP is great as a reporter protein because of the fact that it has no effect on the host once it is introduced to its cellular environment of the host. Due to this ability, no external visualization stain, ATP, or cofactors are needed. With regards to pollutant levels, the fluorescence was measured in order to gauge the effect that the pollutants were have on the host cell. The cellular density of the host cell was also measured. Results from the study conducted by Song, Kim, & Seo (2016) showed that there was a decrease in both fluorescence and cellular density as pollutant levels increased. This was indicative of the fact that cellular activity had decreased. More research into this specific application in order to determine the mechanism by which GFP acts as a pollutant marker.^[39] Similar results have been observed in zebrafish because zebrafish that were injected with GFP were approximately twenty times more susceptible to recognize cellular stresses than zebrafish that were not injected with GFP.^[40]

Advantages[edit]

The biggest advantage of GFP is that it can be heritable, depending on how it was introduced, allowing for continued study of cells and tissues it is expressed in. Visualizing GFP is noninvasive, requiring only illumination with blue light. GFP alone does not interfere with biological processes, but when fused to proteins of interest, careful design of linkers is required to maintain the function of the protein of interest. Moreover, if used with a monomer it is able to diffuse readily throughout cells.^[41]

Fluorescence microscopy[edit]

The availability of GFP and its derivatives has thoroughly redefined fluorescence microscopy and the way it is used in cell biology and other biological disciplines.^[42] While most small fluorescent molecules such as FITC (fluorescein isothiocyanate) are strongly phototoxic when used in live cells, fluorescent proteins such as GFP are usually much less harmful when illuminated in living cells. This has triggered the development of highly automated live-cell fluorescence microscopy systems, which can be used to observe cells over time expressing one or more proteins tagged with fluorescent proteins. For example, GFP had been widely used in labelling the spermatozoa of various organisms for identification purposes as in Drosophila melanogaster, where expression of GFP can be used as a marker for a particular characteristic. GFP can also be expressed in different structures enabling morphological distinction. In such cases, the gene for the production of GFP is incorporated into the genome of the organism in the region of the DNA that codes for the target proteins and that is controlled by the same regulatory sequence; that is, the gene's regulatory sequence now controls the production of GFP, in addition to the tagged protein(s). In cells where the gene is expressed, and the tagged proteins are produced, GFP is produced at the same time. Thus, only those cells in which the tagged gene is expressed, or the target proteins are produced, will fluoresce when observed under fluorescence microscopy. Analysis of such time lapse movies has redefined the understanding of many biological processes including protein folding, protein transport, and RNA dynamics, which in the past had been studied using fixed (i.e., dead) material. Obtained data are also used to calibrate mathematical models of intracellular systems and to estimate rates of gene expression.^[43] Similarly, GFP can be used as an indicator of protein expression in heterologous systems. In this scenario, fusion proteins containing GFP are introduced indirectly, using RNA of the construct, or directly, with the tagged protein itself. This method is useful for studying structural and functional characteristics of the tagged protein on a macromolecular or single-molecule scale with fluorescence microscopy.

The Vertico SMI microscope using the SPDM Phymod technology uses the so-called 'reversible photobleaching' effect of fluorescent dyes like GFP and its derivatives to localize them as single molecules in an optical resolution of 10 nm. This can also be performed as a co-localization of two GFP derivatives (2CLM).^[44]

Another powerful use of GFP is to express the protein in small sets of specific cells. This allows researchers to optically detect specific types of cells in vitro (in a dish), or even in vivo (in the living organism).^[45] Genetically combining several spectral variants of GFP is a useful trick for the analysis of brain circuitry (Brainbow).^[46] Other interesting uses of fluorescent proteins in the literature include using FPs as sensors of neuronmembrane potential,^[47] tracking of AMPA receptors on cell membranes,^[48]viral entry and the infection of individual influenza viruses and lentiviral viruses,^[49][50] etc.

It has also been found that new lines of transgenic GFP rats can be relevant for gene therapy as well as regenerative medicine.^[51] By using 'high-expresser' GFP, transgenic rats display high expression in most tissues, and many cells that have not been characterized or have been only poorly characterized in previous GFP-transgenic rats.

GFP has been shown to be useful in cryobiology as a viability assay. Correlation of viability as measured by trypan blue assays were 0.97.^[52] Another application is the use of GFP co-transfection as internal control for transfection efficiency in mammalian cells.^[53]

A novel possible use of GFP includes using it as a sensitive monitor of intracellular processes via an eGFP laser system made out of a human embryonic kidney cell line. The first engineered living laser is made by an eGFP expressing cell inside a reflective optical cavity and hitting it with pulses of blue light. At a certain pulse threshold, the eGFP's optical output becomes brighter and completely uniform in color of pure green with a wavelength of 516 nm. Before being emitted as laser light, the light bounces back and forth within the resonator cavity and passes the cell numerous times. By studying the changes in optical activity, researchers may better understand cellular processes.^[54][55]

GFP is used widely in cancer research to label and track cancer cells. GFP-labelled cancer cells have been used to model metastasis, the process by which cancer cells spread to distant organs.^[56]

Split GFP[edit]

GFP can be used to analyse the colocalization of proteins. This is achieved by 'splitting' the protein into two fragments which are able to self-assemble, and then fusing each of these to the two proteins of interest. Alone, these incomplete GFP fragments are unable to fluoresce. However, if the two proteins of interest colocalize, then the two GFP fragments assemble together to form a GFP-like structure which is able to fluoresce. Therefore, by measuring the level of fluorescence it is possible to determine whether the two proteins of interest colocalize.^[57]

TECHNOLOGIES FOR CAREER TRANSITION. Developing managerial skills in engineers and scientists pdf writer jobs. TRANSFORMATION OF TECHNOLOGISTS INTO MANAGERS. Technologies for Management. Issues in Career Transition. Preventing Managerial Failure.

Macro-photography[edit]

Macro-scale biological processes, such as the spread of virus infections, can be followed using GFP labeling.^[58] In the past, mutagenic ultra violet light (UV) has been used to illuminate living organisms (e.g., see^[59]) to detect and photograph the GFP expression. Recently, a technique using non-mutagenic LED lights^[60] have been developed for macro-photography.^[61] The technique uses an epifluorescence camera attachment^[62] based on the same principle used in the construction of epifluorescence microscopes.

Transgenic pets[edit]

Alba, a green-fluorescent rabbit, was created by a French laboratory commissioned by Eduardo Kac using GFP for purposes of art and social commentary.^[63] The US company Yorktown Technologies markets to aquarium shops green fluorescent zebrafish (GloFish) that were initially developed to detect pollution in waterways. NeonPets, a US-based company has marketed green fluorescent mice to the pet industry as NeonMice.^[64] Green fluorescent pigs, known as Noels, were bred by a group of researchers led by Wu Shinn-Chih at the Department of Animal Science and Technology at National Taiwan University.^[65] A Japanese-American Team created green-fluorescent cats as proof of concept to use them potentially as model organisms for diseases, particularly HIV.^[66] In 2009 a South Korean team from Seoul National University bred the first transgenic beagles with fibroblast cells from sea anemones. The dogs give off a red fluorescent light, and they are meant to allow scientists to study the genes that cause human diseases like narcolepsy and blindness.^[67]

Art[edit]

Julian Voss-Andreae, a German-born artist specializing in 'protein sculptures,'^[68] created sculptures based on the structure of GFP, including the 1.70 m (5'6') tall 'Green Fluorescent Protein' (2004)^[69] and the 1.40 m (4'7') tall 'Steel Jellyfish' (2006). The latter sculpture is located at the place of GFP's discovery by Shimomura in 1962, the University of Washington's Friday Harbor Laboratories.^[70]

See also[edit]

References[edit]

1. ^Ormö M, Cubitt AB, Kallio K, Gross LA, Tsien RY, Remington SJ (September 1996). 'Crystal structure of the Aequorea victoria green fluorescent protein'. Science. 273 (5280): 1392–5. Bibcode:1996Sci..273.1392O. doi:10.1126/science.273.5280.1392. PMID8703075.
2. ^Prendergast FG, Mann KG (Aug 1978). 'Chemical and physical properties of aequorin and the green fluorescent protein isolated from Aequorea forskålea'. Biochemistry. 17 (17): 3448–53. doi:10.1021/bi00610a004. PMID28749.
3. ^ ^abcdTsien RY (1998). 'The green fluorescent protein'(PDF). Annual Review of Biochemistry. 67: 509–44. doi:10.1146/annurev.biochem.67.1.509. PMID9759496.
4. ^Stepanenko OV, Verkhusha VV, Kuznetsova IM, Uversky VN, Turoverov KK (Aug 2008). 'Fluorescent proteins as biomarkers and biosensors: throwing color lights on molecular and cellular processes'. Current Protein & Peptide Science. 9 (4): 338–69. doi:10.2174/138920308785132668. PMC2904242. PMID18691124.
5. ^Phillips GJ (Oct 2001). 'Green fluorescent protein--a bright idea for the study of bacterial protein localization'. FEMS Microbiology Letters. 204 (1): 9–18. doi:10.1016/S0378-1097(01)00358-5. PMID11682170.
6. ^Shimomura O, Johnson FH, Saiga Y (Jun 1962). 'Extraction, purification and properties of aequorin, a bioluminescent protein from the luminous hydromedusan, Aequorea'. Journal of Cellular and Comparative Physiology. 59 (3): 223–39. doi:10.1002/jcp.1030590302. PMID13911999.
7. ^Morise H, Shimomura O, Johnson FH, Winant J (Jun 1974). 'Intermolecular energy transfer in the bioluminescent system of Aequorea'. Biochemistry. 13 (12): 2656–62. doi:10.1021/bi00709a028. PMID4151620.
8. ^Prasher DC, Eckenrode VK, Ward WW, Prendergast FG, Cormier MJ (Feb 1992). 'Primary structure of the Aequorea victoria green-fluorescent protein'. Gene. 111 (2): 229–33. doi:10.1016/0378-1119(92)90691-H. PMID1347277.
9. ^Chalfie M, Tu Y, Euskirchen G, Ward WW, Prasher DC (Feb 1994). 'Green fluorescent protein as a marker for gene expression'. Science. 263 (5148): 802–5. Bibcode:1994Sci..263.802C. doi:10.1126/science.8303295. PMID8303295.
10. ^Inouye S, Tsuji FI (Mar 1994). 'Aequorea green fluorescent protein. Expression of the gene and fluorescence characteristics of the recombinant protein'. FEBS Letters. 341 (2–3): 277–80. doi:10.1016/0014-5793(94)80472-9. PMID8137953.
11. ^ ^abcOrmö M, Cubitt AB, Kallio K, Gross LA, Tsien RY, Remington SJ (Sep 1996). 'Crystal structure of the Aequorea victoria green fluorescent protein'. Science. 273 (5280): 1392–5. Bibcode:1996Sci..273.1392O. doi:10.1126/science.273.5280.1392. PMID8703075.
12. ^ ^abcYang F, Moss LG, Phillips GN (Oct 1996). 'The molecular structure of green fluorescent protein'(PDF). Nature Biotechnology. 14 (10): 1246–51. doi:10.1038/nbt1096-1246. hdl:1911/19233. PMID9631087.
13. ^ ^abcBrejc, K.; Sixma, T. K.; Kitts, P. A.; Kain, S. R.; Tsien, R. Y.; Ormö, M.; Remington, S. J. Structural basis for dual excitation and photoisomerization of the Aequorea victoria green fluorescent protein. Proc. Natl. Acad. Sci. U. S. A.1997, 94 (6), 2306-2311.
14. ^Shaner NC, Steinbach PA, Tsien RY (Dec 2005). 'A guide to choosing fluorescent proteins'(PDF). Nature Methods. 2 (12): 905–9. doi:10.1038/nmeth819. PMID16299475.
15. ^Wilhelmsson and Tor (2016). Fluorescent Analogs of Biomolecular Building Blocks: Design and Applications. New Jersey: Wiley. ISBN978-1-118-17586-6.
16. ^Heim R, Cubitt AB, Tsien RY (Feb 1995). 'Improved green fluorescence'(PDF). Nature. 373 (6516): 663–4. Bibcode:1995Natur.373.663H. doi:10.1038/373663b0. PMID7854443.
17. ^US patent 6172188, Thastrup O, Tullin S, Kongsbak Poulsen L, Bjørn S, 'Fluorescent Proteins', published 2001-01-09
18. ^Cormack BP, Valdivia RH, Falkow S (1996). 'FACS-optimized mutants of the green fluorescent protein (GFP)'. Gene. 173 (1 Spec No): 33–38. doi:10.1016/0378-1119(95)00685-0. PMID8707053.
19. ^McRae SR, Brown CL, Bushell GR (May 2005). 'Rapid purification of EGFP, EYFP, and ECFP with high yield and purity'. Protein Expression and Purification. 41 (1): 121–127. doi:10.1016/j.pep.2004.12.030. PMID15802229.
20. ^Pédelacq JD, Cabantous S, Tran T, Terwilliger TC, Waldo GS (Jan 2006). 'Engineering and characterization of a superfolder green fluorescent protein'. Nature Biotechnology. 24 (1): 79–88. doi:10.1038/nbt1172. PMID16369541.
21. ^Barondeau DP, Kassmann CJ, Tainer JA, Getzoff ED (Apr 2002). 'Structural chemistry of a green fluorescent protein Zn biosensor'. Journal of the American Chemical Society. 124 (14): 3522–3524. doi:10.1021/ja0176954. PMID11929238.
22. ^Lelimousin M, Noirclerc-Savoye M, Lazareno-Saez C, Paetzold B, Le Vot S, Chazal R, Macheboeuf P, Field MJ, Bourgeois D, Royant A (Oct 2009). 'Intrinsic dynamics in ECFP and Cerulean control fluorescence quantum yield'. Biochemistry. 48 (42): 10038–10046. doi:10.1021/bi901093w. PMID19754158.
23. ^Goedhart J, von Stetten D, Noirclerc-Savoye M, Lelimousin M, Joosen L, Hink MA, van Weeren L, Gadella TW, Royant A (2012). 'Structure-guided evolution of cyan fluorescent proteins towards a quantum yield of 93%'. Nature Communications. 3: 751. Bibcode:2012NatCo..3.751G. doi:10.1038/ncomms1738. PMC3316892. PMID22434194.
24. ^Miesenböck G, De Angelis DA, Rothman JE (Jul 1998). 'Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins'. Nature. 394 (6689): 192–5. Bibcode:1998Natur.394.192M. doi:10.1038/28190. PMID9671304.
25. ^Hanson GT, Aggeler R, Oglesbee D, Cannon M, Capaldi RA, Tsien RY, Remington SJ (Mar 2004). 'Investigating mitochondrial redox potential with redox-sensitive green fluorescent protein indicators'. The Journal of Biological Chemistry. 279 (13): 13044–53. doi:10.1074/jbc.M312846200. PMID14722062.
26. ^Zacharias DA, Violin JD, Newton AC, Tsien RY (May 2002). 'Partitioning of lipid-modified monomeric GFPs into membrane microdomains of live cells'. Science. 296 (5569): 913–16. Bibcode:2002Sci..296.913Z. doi:10.1126/science.1068539. PMID11988576.
27. ^Chudakov DM, Belousov VV, Zaraisky AG, Novoselov VV, Staroverov DB, Zorov DB, Lukyanov S, Lukyanov KA (February 2003). 'Kindling fluorescent proteins for precise in vivo photolabeling'. Nature Biotechnology. 21 (2): 191–4. doi:10.1038/nbt778. PMID12524551.
28. ^Wiens MD, Shen Y, Li X, Salem MA, Smisdom N, Zhang W, Brown A, Campbell RE (December 2016). 'A Tandem Green-Red Heterodimeric Fluorescent Protein with High FRET Efficiency'. ChemBioChem. 17 (24): 2361–2367. doi:10.1002/cbic.201600492. PMID27781394.
29. ^Drepper, T., Eggert, T., Circolone, F., Heck, A., Krauss, U., Guterl, J. K., Wendorff, M., Losi, A., Gärtner, W., Jaeger, K. E. (2007). 'Reporter proteins for in vivo fluorescence without oxygen'. Nat Biotechnol. 25 (4): 443–445. doi:10.1038/nbt1293. PMID17351616.CS1 maint: multiple names: authors list (link)
30. ^ ^abRodriguez EA, Tran GN, Gross LA, Crisp JL, Shu X, Lin JY, Tsien RY (September 2016). 'A far-red fluorescent protein evolved from a cyanobacterial phycobiliprotein'. Nature Methods. 13 (9): 763–9. doi:10.1038/nmeth.3935. PMC5007177. PMID27479328.
31. ^Tsien RY (1998-01-01). 'The green fluorescent protein'. Annual Review of Biochemistry. 67 (1): 509–44. doi:10.1146/annurev.biochem.67.1.509. PMID9759496.
32. ^Rodriguez EA, Campbell RE, Lin JY, Lin MZ, Miyawaki A, Palmer AE, Shu X, Zhang J, Tsien RY (February 2017). 'The Growing and Glowing Toolbox of Fluorescent and Photoactive Proteins'. Trends in Biochemical Sciences. 42 (2): 111–129. doi:10.1016/j.tibs.2016.09.010. PMC5272834. PMID27814948.
33. ^Bokman SH, Ward WW (1982). 'Reversible denaturation of Aequorea green-fluorescent protein: physical separation and characterization of the renatured protein'. Biochemistry. 21 (19): 4535–4540. doi:10.1021/bi00262a003.
34. ^Pouwels LJ, Zhang L, Chan NH, Dorrestein PC, Wachter RM (Sep 2008). 'Kinetic isotope effect studies on the de novo rate of chromophore formation in fast- and slow-maturing GFP variants'. Biochemistry. 47 (38): 10111–22. doi:10.1021/bi8007164. PMC2643082. PMID18759496.
35. ^Chudakov DM, Matz MV, Lukyanov S, Lukyanov KA (Jul 2010). 'Fluorescent proteins and their applications in imaging living cells and tissues'. Physiological Reviews. 90 (3): 1103–63. doi:10.1152/physrev.00038.2009. PMID20664080.
36. ^Stepaneko, O. V.; Verkhusha, V. V.; Shavlovsky, M. M.; Kuznetsova, I. M.; Uversky, V. N.; Turoverov, K. K. Understanding the role of Arg96 in structure and stability of green fluorescent protein. Proteins: Struct., Funct., Bioinf.1999, 73 (3), 539-551.
37. ^Jugder BE, Welch J, Braidy N, Marquis CP (2016-07-26). 'Construction and use of a Cupriavidus necator H16 soluble hydrogenase promoter (PSH) fusion to gfp (green fluorescent protein)'. PeerJ. 4: e2269. doi:10.7717/peerj.2269. PMC4974937. PMID27547572.
38. ^Arun KH, Kaul CL, Ramarao P (2005). 'Green fluorescent proteins in receptor research: an emerging tool for drug discovery'. Journal of Pharmacological and Toxicological Methods. 51 (1): 1–23. doi:10.1016/j.vascn.2004.07.006. PMID15596111.
39. ^Song YH, Kim CS, Seo JH (April 2016). 'Noninvasive monitoring of environmental toxicity through green fluorescent protein expressing Escherichia coli'. Korean Journal of Chemical Engineering. 33 (4): 1331–6. doi:10.1007/s11814-015-0253-1.
40. ^Pan Y, Leifert A, Graf M, Schiefer F, Thoröe-Boveleth S, Broda J, Halloran MC, Hollert H, Laaf D, Simon U, Jahnen-Dechent W (March 2013). 'High-sensitivity real-time analysis of nanoparticle toxicity in green fluorescent protein-expressing zebrafish'. Small. Weinheim an Der Bergstrasse, Germany. 9 (6): 863–9. doi:10.1002/smll.201201173. PMID23143852.
41. ^Chalfie M (Jun 2009). 'GFP: Lighting up life'. Proceedings of the National Academy of Sciences of the United States of America. 106 (25): 10073–10080. Bibcode:2009PNAS.10610073C. doi:10.1073/pnas.0904061106. PMC2700921. PMID19553219.
42. ^Yuste R (Dec 2005). 'Fluorescence microscopy today'. Nature Methods. 2 (12): 902–4. doi:10.1038/nmeth1205-902. PMID16299474.
43. ^Komorowski M, Finkenstädt B, Rand D (Jun 2010). 'Using a single fluorescent reporter gene to infer half-life of extrinsic noise and other parameters of gene expression'. Biophysical Journal. 98 (12): 2759–2769. Bibcode:2010BpJ..98.2759K. doi:10.1016/j.bpj.2010.03.032. PMC2884236. PMID20550887.
44. ^Gunkel M, Erdel F, Rippe K, Lemmer P, Kaufmann R, Hörmann C, Amberger R, Cremer C (Jun 2009). 'Dual color localization microscopy of cellular nanostructures'. Biotechnology Journal. 4 (6): 927–38. doi:10.1002/biot.200900005. PMID19548231.
45. ^Chudakov DM, Lukyanov S, Lukyanov KA (Dec 2005). 'Fluorescent proteins as a toolkit for in vivo imaging'. Trends in Biotechnology. 23 (12): 605–13. doi:10.1016/j.tibtech.2005.10.005. PMID16269193.
46. ^Livet J, Weissman TA, Kang H, Draft RW, Lu J, Bennis RA, Sanes JR, Lichtman JW (Nov 2007). 'Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous system'. Nature. 450 (7166): 56–62. Bibcode:2007Natur.450..56L. doi:10.1038/nature06293. PMID17972876.
47. ^Baker BJ, Mutoh H, Dimitrov D, Akemann W, Perron A, Iwamoto Y, Jin L, Cohen LB, Isacoff EY, Pieribone VA, Hughes T, Knöpfel T (Aug 2008). 'Genetically encoded fluorescent sensors of membrane potential'. Brain Cell Biology. 36 (1–4): 53–67. doi:10.1007/s11068-008-9026-7. PMC2775812. PMID18679801.
48. ^Adesnik H, Nicoll RA, England PM (Dec 2005). 'Photoinactivation of native AMPA receptors reveals their real-time trafficking'. Neuron. 48 (6): 977–85. doi:10.1016/j.neuron.2005.11.030. PMID16364901.
49. ^Lakadamyali M, Rust MJ, Babcock HP, Zhuang X (Aug 2003). 'Visualizing infection of individual influenza viruses'. Proceedings of the National Academy of Sciences of the United States of America. 100 (16): 9280–5. Bibcode:2003PNAS.100.9280L. doi:10.1073/pnas.0832269100. PMC170909. PMID12883000.
50. ^Joo KI, Wang P (Oct 2008). 'Visualization of targeted transduction by engineered lentiviral vectors'. Gene Therapy. 15 (20): 1384–96. doi:10.1038/gt.2008.87. PMC2575058. PMID18480844.
51. ^Remy S, Tesson L, Usal C, Menoret S, Bonnamain V, Nerriere-Daguin V, Rossignol J, Boyer C, Nguyen TH, Naveilhan P, Lescaudron L, Anegon I (Oct 2010). 'New lines of GFP transgenic rats relevant for regenerative medicine and gene therapy'. Transgenic Research. 19 (5): 745–63. doi:10.1007/s11248-009-9352-2. PMID20094912.
52. ^Elliott G, McGrath J, Crockett-Torabi E (Jun 2000). 'Green fluorescent protein: A novel viability assay for cryobiological applications'. Cryobiology. 40 (4): 360–369. doi:10.1006/cryo.2000.2258. PMID10924267.
53. ^Fakhrudin N, Ladurner A, Atanasov AG, Heiss EH, Baumgartner L, Markt P, Schuster D, Ellmerer EP, Wolber G, Rollinger JM, Stuppner H, Dirsch VM (Apr 2010). 'Computer-aided discovery, validation, and mechanistic characterization of novel neolignan activators of peroxisome proliferator-activated receptor gamma'. Molecular Pharmacology. 77 (4): 559–66. doi:10.1124/mol.109.062141. PMC3523390. PMID20064974.
54. ^Gather MC, Yun SH (2011). 'Single-cell biological lasers'. Nature Photonics. 5 (7): 406–410. Bibcode:2011NaPho..5.406G. doi:10.1038/nphoton.2011.99.
55. ^Matson J (2011). 'Green Fluorescent Protein Makes for Living Lasers'. Scientific American. Retrieved 2011-06-13.
56. ^Kouros-Mehr H, Bechis SK, Slorach EM, Littlepage LE, Egeblad M, Ewald AJ, Pai SY, Ho IC, Werb Z (Feb 2008). 'GATA-3 links tumor differentiation and dissemination in a luminal breast cancer model'. Cancer Cell. 13 (2): 141–52. doi:10.1016/j.ccr.2008.01.011. PMC2262951. PMID18242514.
57. ^Cabantous S, Terwilliger TC, Waldo GS (January 2005). 'Protein tagging and detection with engineered self-assembling fragments of green fluorescent protein'. Nature Biotechnology. 23 (1): 102–7. doi:10.1038/nbt1044. PMID15580262.
58. ^Rodman MK, Yadav NS, Artus NN (2002-09-01). 'Progression of geminivirus-induced transgene silencing is associated with transgene methylation'. New Phytologist. 155 (3): 461–468. doi:10.1046/j.1469-8137.2002.00467.x.
59. ^Zhu YJ, Agbayani R, Moore PH (Apr 2004). 'Green fluorescent protein as a visual selection marker for papaya (Carica papaya L.) transformation'. Plant Cell Reports. 22 (9): 660–7. doi:10.1007/s00299-004-0755-5. PMID14749892.
60. ^Niwa Y, Hirano T, Yoshimoto K, Shimizu M, Kobayashi H (1999). 'Non-invasive quantitative detection and applications of non-toxic, S65T-type green fluorescent protein in living plants'. The Plant Journal. 18 (4): 455–63. doi:10.1046/j.1365-313X.1999.00464.x. PMID10406127.
61. ^Baker SS, Vidican CB, Cameron DS, Greib HG, Jarocki CC, Setaputri AW, Spicuzza CH, Burr AA, Waqas MA, Tolbert DA (2012-01-01). 'An epifluorescent attachment improves whole-plant digital photography of Arabidopsis thaliana expressing red-shifted green fluorescent protein'. AoB PLANTS. 2012: pls003. doi:10.1093/aobpla/pls003. PMC3296078. PMID22479674.
62. ^'PlantEdDL - Using SRL digital cameras in quantitative investigations of plants expressing green fluorescent protein (GFP)'. planted.botany.org. Retrieved 2016-03-23.
63. ^Eduardo Kac. 'GFP Bunny'.
64. ^'Glow-In-The Dark NeonMice'. Retrieved August 30, 2016.Cite uses deprecated parameter dead-url= (help)^{[dead link]}
65. ^Scientists in Taiwan breed fluorescent green pigs
66. ^Wongsrikeao P, Saenz D, Rinkoski T, Otoi T, Poeschla E (2011). 'Antiviral restriction factor transgenesis in the domestic cat'. Nature Methods. 8 (10): 853–9. doi:10.1038/nmeth.1703. PMC4006694. PMID21909101.
67. ^'Fluorescent puppy is world's first transgenic dog'.
68. ^Voss-Andreae J (2005). 'Protein Sculptures: Life's Building Blocks Inspire Art'. Leonardo. 38: 41–45. doi:10.1162/leon.2005.38.1.41.
69. ^Pawlak A (2005). 'Inspirierende Proteine'. Physik Journal. 4: 12.
70. ^'Julian Voss-Andreae Sculpture'. Retrieved 2007-06-14.

Further reading[edit]

• Pieribone V, Gruber D (2006). Aglow in the Dark: The Revolutionary Science of Biofluorescence. Cambridge: Belknap Press. ISBN978-0-674-01921-8. OCLC60321612. Popular science book describing history and discovery of GFP
• Zimmer M (2005). Glowing Genes: A Revolution In Biotechnology. Buffalo, NY: Prometheus Books. ISBN978-1-59102-253-4. OCLC56614624.

External links[edit]

• Interactive Java applet demonstrating the chemistry behind the formation of the GFP chromophore
• Green Fluorescent Protein Chem Soc Rev themed issue dedicated to the 2008 Nobel Prize winners in Chemistry, Professors Osamu Shimomura, Martin Chalfie and Roger Y. Tsien
• Molecule of the Month, June 2003: an illustrated overview of GFP by David Goodsell.
• Molecule of the Month, June 2014: an illustrated overview of GFP-like variants by David Goodsell.
• Green Fluorescent Protein on FPbase