KW - Green fluorescent protein (GFP)
The underlying concept was that synthetic GFP chromophore analogues are not fluorescent primarily because of free rotation about an aryl−alkene bond (Figure 1b).
GFP Proton Shuttle Reaction IRC

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Green fluorescent protein - Wikipedia
1 The chromophore within GFP is formed from a Ser-Tyr-Gly tripeptide in the primary protein structure by post-translational cyclodehydration and subsequent oxidation to yield the 4-( p -hydroxybenzylidene)imidazolinone (HO-BDI) moiety.
The green fluorescent protein ..
AB - Green fluorescent protein (GFP) has a chromophore that forms autocatalytically within the folded protein. Although many studies have focused on the precise mechanism of chromophore maturation, little is known about the kinetics of de novo chromophore maturation. Here we present a simple and efficient method for examining the de novo kinetics. GFP with an immature chromophore was synthesized in a reconstituted cell-free protein synthesis system under anaerobic conditions. Chromophore maturation was initiated by rapid dilution in an air-saturated maturation buffer, and the time course of fluorescence development was monitored. Comparison of the de novo maturation rates in various GFP variants revealed that some folding mutations near the chromophore promoted rapid chromophore maturation and that the accumulation of mutations could reduce the maturation rate. Our method will contribute to the design of rapidly maturing fluorescent proteins with improved characteristics for real-time monitoring of cellular events.

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