The green fluorescent protein is a protein composed of 238 amino acid residues that exhibits bright green fluorescence when exposed to light in the blue to ultraviolet range. Although many other marine organisms have similar green fluorescent proteins, GFP traditionally refers to the protein first isolated from the jellyfish Aequorea 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, in the lower green portion of the visible spectrum; the fluorescence quantum yield of GFP is 0.79. The GFP from the sea pansy 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. In cell and molecular biology, the GFP gene is used as a reporter of expression, it has been used in modified forms to make biosensors, 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, maintained in their genome and that of their offspring. To date, GFP has been expressed in many species, including bacteria, fungi and mammals, including in human cells. Scientists Roger Y. Tsien, Osamu Shimomura, Martin Chalfie were awarded the 2008 Nobel Prize in Chemistry on 10 October 2008 for their discovery and development of the green fluorescent protein. In the 1960s and 1970s, GFP, along with the separate luminescent protein aequorin, was first purified from Aequorea victoria and its properties studied by Osamu Shimomura. 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. 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; 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. Frederick Tsuji's lab independently reported the expression of the recombinant protein one month later. 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. One month the Phillips group independently reported the wild-type GFP structure in Nature Biotechnology; 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, guanidinium chloride treatments, drastic temperature changes. Due to the potential for widespread usage and the evolving needs of researchers, many different mutants of GFP have been engineered; the first major improvement was a single point mutation reported in 1995 in Nature by Roger Tsien. This mutation improved the spectral characteristics of GFP, resulting in increased fluorescence, a shift of the major excitation peak to 488 nm, with the peak emission kept at 509 nm; this matched the spectral characteristics of available FITC filter sets, increasing the practicality of use by the general researcher. A 37 °C folding efficiency point mutant to this scaffold, yielding enhanced GFP, was discovered in 1995 by the laboratories of Thastrup and Falkow. EGFP allowed the practical use of GFPs in mammalian cells.
EGFP has an extinction coefficient of 55,000 M−1cm−1. The fluorescence quantum yield of EGFP is 0.60. The relative brightness, expressed as ε•QY, is 33,000 M−1cm−1. Superfolder GFP, a series of mutations that allow GFP to fold and mature when fused to poorly folding peptides, was reported in 2006. Many other mutations have been made, including color mutants. BFP derivatives 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 that preferentially binds Cu has been developed. BFPms1 have several important mutations including and the BFP chromophore,Y145F for higher quantum yield, H148G for creating a hole into the beta-barrel and several other mutations that increase solubility. Zn binding increases fluorescence intensity, while Cu binding quenches fluorescence and shifts the absorbance maximum from 379 to 444 nm. Therefore, they can be used as Zn biosensor.
Nicolas Alejandro Cinalli is a former Argentine professional footballer who plays as a goalkeeper. Born in Rosario, Cinalli played youth football with Colegio Salesiano San Jose and Central Córdoba, before turning professional with the latter in 1999. After one season he moved to Italy, spending a number of years in the lower leagues with teams such as Virtus Entella, Ragusa, Potenza and played one season in the Italian Serie "A" with Perugia before moving to the Netherlands Eredivisie in 2007, to sign for De Graafschap. Cinalli spent two seasons with De Graafschap, he ended his career in Argentina playing for Atletico Policial, El Porvenir del Norte and Sportivo Las Parejas in the Torneo Argentino "B"
FAP Captain David Abensur Rengifo International Airport is an airport serving the city of Pucallpa in the Ucayali Region of Peru. It is operated by Aeropuertos del Perú, S. A. Captain Rengifo Airport is the main airport serving the Ucayali Region; the Pucallpa VOR-DME is located on the field. North American Flote and AirMajoro provide charter flights to amazon towns in Peru. On 23 August 2005, TANS Perú Flight 204, a Boeing 737, crashed short of the runway at Pucallpa Airport. Out of the 98 passengers and crew on board, 40 were killed. Transport in Peru List of airports in Peru Airport information for SPCL at World Aero Data. Data current as of October 2006. CORPAC official site SkyVector Aeronautical Charts OurAirports - Pucallpa OpenStreetMap - Pucallpa