Green-fluorescent protein from the bioluminescent jellyfish Clytia gregaria: cDNA cloning, expression, and characterization of novel recombinant protein.
The bioluminescent systems of many marine organisms are comprised of two proteins--the Ca(2+)-regulated photoprotein and green-fluorescent protein (GFP). This work reports the cloning of the full-size cDNA encoding GFP (cgreGFP) from jellyfish Clytia gregaria, its expression and properties of the recombinant protein. The overall degree of identity between the amino acid sequence of the novel cgreGFP and the sequence of GFP (avGFP) from Aequorea victoria is 42% (similarity--64%) despite these GFPs originating from jellyfish that both belong to the same class, Hydrozoa. However although the degree of identity is low, three residues, Ser-Tyr-Gly, which form the chromophore are identical in both GFPs. The cgreGFP displayed two absorption peaks at 278 and 485 nm, and the fluorescence maximum at 500 nm. The fluorescence quantum yield was determined to be 0.86, the brightness to be 54 mM(-1) cm(-1). For the first time we have also demonstrated an efficient radiationless energy transfer in vitro between clytin and cgreGFP in solution at micromolar concentrations. The cgreGFP may be a useful intracellular fluorescent marker, as it was able to be expressed in mammalian cells.
Markova
Svetlana V
SV
Photobiology Lab, Institute of Biophysics, Russian Academy of Sciences Siberian Branch, Akademgorodok 50, Bldg. 50, Krasnoyarsk, 660036, Russia.
Burakova
Ludmila P
LP
Frank
Ludmila A
LA
Golz
Stefan
S
Korostileva
Kseniya A
KA
Vysotski
Eugene S
ES
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D029941:Fish Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008164:Luminescent Proteins / Q000737:chemistry; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D037401:Scyphozoa / Q000378:metabolism*; D016415:Sequence Alignment; D013050:Spectrometry, Fluorescence
D029941:Fish Proteins; D008164:Luminescent Proteins; D011994:Recombinant Proteins; C046165:phialidin; D049452:Green Fluorescent Proteins