Timeline Stories – Page 3 – Green Fluorescent Protein

First crystal structures of wild-type and enhanced GFP. Tsien designed T203Y mutant based on crystal structure of S65T GFP. It is yellow fluorescent.

References: M Ormo, AB Cubitt, K Kallio, LA Gross, RY Tsien, SJ Remington: Crystal structure of the Aequorea victoria green fluorescent protein. Science 273 (1996) 1392-95. F Yang, LG Moss,…

Posttranslational oxidation of S65T GFP four times faster than wt-GFP. The origin of enhanced GFP (EGFP).

Reference: R Heim, A Cubitt, RY Tsien: Improved green fluorescene. Nature 373 (1995) 663-64.

GFP expressed in E. coli and C. elegans. Mechanism for chromophore formation. First new color (blue) and oxygen dependence reported. Possibility of use in FRET experiments mentioned.

References: M Chalfie, Y Tu, G Euskirchen, WW Ward, DC Prasher: Green fluorescent protein as a marker for gene expression. Science 263 (1994) 802-05. S Inouye, F Tsuji: Evidence for…

Structure of GFP chromophore confirmed, flanking amino acid residues corrected from Shimomura’s 1979 structure. Still “apoGFP”

CW Cody, DC Prasher, WM Westler, FG Pendergast, WW Ward: Chemical Structure of the hexapeptide chromophore of the Aequorea Green fluorescent protein. Biochemistry 32 (1993) 1212-18.

GFP cloned. “These results will enable us to construct an expression vector for the preparation of non-fluorescent apoGFP.”

Reference: DC Prasher, VK Eckenrode, WW Ward, FG Pendergast, MJ Cormier: Primary structure of the Aequorea victorea green fluorescent protein. Gene 111 (1992) 229-33.

Prasher clones and expresses aequorin

Reference: D Prasher, RO McCann, MJ Cormier: Cloning and Expression of the Cdna Coding for Aequorin, a Bioluminescent Calcium-Binding Protein. Biochemical and Biophysical Research Communications 126 (1985) 1259-68.

Shimomura characterized structure of chromophore

O Shimomura: Structure of the chromophore of Aequorea green fluorescent protein. FEBS Letters 104 (1979) 220-22.

Intermolecular energy transfer between aequorin and GFP in jellyfish and when co-absorbed on Sephadex column.

References: JG Morin, JW Hastings: Energy Transfer in a bioluminescent system. J. Cell Physiol. 77 (1971) 313-18. H Morise, O Shimomura, FH Johnson, J Winant: Intermolecular Energy Transfer in Bioluminescent…

“Green protein” named green fluorescent protein.

Reference: JW Hasting, JG Morin: Comparative biochemistry of calcium-activated photoproteins from the ctenophore, Mnemiopsis, and the coelenterates Aequorea, Obelia, Pelagia and Renilla. Biol. Bull. 137 (1969) 402.

GFP identified as protein, extracted from 10,000 jellyfish – “a protein giving solutions that look slightly greenish in sunlight though only yellowish under tungsten lights, and exhibiting a very bright, greenish fluorescence in the ultraviolet of a Mineralite, has also been isolated from squeezates.” Called “green protein.”

Reference: O Shimomura, FH Johnson, Y Saiga: Extraction, purification and properties of aequorin, a bioluminescent protein from the luminous hydromedusan, Aequorea. J. Cell. Comp. Physiol. 59 (1962) 223-29.