Not yet tested in other applications.
Optimal dilutions/concentrations should be determined by the end user.
Function: Energy-transfer acceptor. Its role is to transduce the blue chemiluminescence of the protein aequorin into green fluorescent light by energy transfer. Fluoresces in vivo upon receiving energy from the Ca2+ -activated photoprotein aequorin.
Subunit structure: Monomer.
Tissue specificity: Photocytes.
Post-translational modification: Contains a chromophore consisting of modified amino acid residues. The chromophore is formed by autocatalytic backbone condensation between Ser-65 and Gly-67, and oxidation of Tyr-66 to didehydrotyrosine. Maturation of the chromophore requires nothing other than molecular oxygen.
Biotechnological use: Green fluorescent protein has been engineered to produce a vast number of variously colored mutants, fusion proteins, and biosensors. Fluorescent proteins and its mutated allelic forms, blue, cyan and yellow have become a useful and ubiquitous tool for making chimeric proteins, where they function as a fluorescent protein tag. Typically they tolerate N- and C-terminal fusion to a broad variety of proteins. They have been expressed in most known cell types and are used as a noninvasive fluorescent marker in living cells and organisms. They enable a wide range of applications where they have functioned as a cell lineage tracer, reporter of gene expression, or as a measure of protein-protein interactions. Can also be used as a molecular thermometer, allowing accurate temperature measurements in fluids. The measurement process relies on the detection of the blinking of GFP using fluorescence correlation spectroscopy.
Sequence similarities: Belongs to the GFP family.
Biophysicochemical properties: Absorption: Abs(max)=395 nm
Exhibits a smaller absorbance peak at 470 nm. The fluorescence emission spectrum peaks at 509 nm with a shoulder at 540 nm.
Following fixation with 1X PBS, 3.5 % formaldehyde, pCMV5-Yes-GFP transfected COS-7 cells were permeabilized with 1X PBS, 0.1 % Triton-X100 (A and B) as described in (McCabe and Berthiaume 1999) and GFP fluorescence was quenched by treatment with 100 mM MES pH 6.5 for 30 minutes (C) (Berthiaume, L.G., unpublished). Cells were then labelled with or without rabbit anti-GFP-FITC at a concentration of 1 ug/ml as described in (McCabe and Berthiaume 1999). A) 200X magnification of COS-7 cells expressing Yes-GFP, B) 1000X magnification of boxed area in A and C) 200X magnification of quenched COS-7 cells expressing Yes-GFP. 100 um bar in A and C; 20?um bar in inset B.
McCabe, J.B. and Berthiaume, L.G. (1999) Functional roles for fatty acylated N-terminal domains in subcellular localization. Mol. Biol. Cell, 10, 3771-3786.