Chemical nature of the light emitter of the Aequorea green fluorescent protein
Chemical nature of the light emitter of the Aequorea green fluorescent protein
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DOI：
10.1073/pnas.93.24.13617
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发表时间：
1996-11-26
影响因子：
11.1
通讯作者：
Tsuji, FI
中科院分区：
文献类型：
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作者：
Niwa, H;Inouye, S;Tsuji, FI
维多利亚Aequorea水母在其保护伞边缘有一种绿色荧光蛋白（GFP, 27 kDa），在动物的生物发光反应中充当最终的光发射器，该蛋白由单个多肽链中的238个氨基酸残基组成，在长紫外光照射下产生绿色荧光（λ (max) = 508 nm）。荧光是由于存在由咪唑酮环组成的发色团，该发色团由三肽- ser (65)- tyr (66)- gly(67)-的翻译后修饰形成。在真核和原核细胞中，绿色荧光蛋白已被广泛用作监测基因表达的报告蛋白，但对荧光产生的化学机制知之甚少。为了更好地理解这一问题，我们研究了带有发色团的绿色荧光蛋白肽片段和合成的发色团模型化合物。结果表明，GFP发色团由咪唑酮环结构组成，发光源为发色团中酚酸阴离子的单线态激发态，发光高度依赖于发色团周围的微环境，而发色团的外亚甲基双键异构化抑制是其有效发光的原因。
The jellyfish Aequorea victoria possesses in the margin of its umbrella a green fluorescent protein (GFP, 27 kDa) that serves as the ultimate light emitter in the bioluminescence reaction of the animal, The protein is made up of 238 amino acid residues in a single polypeptide chain and produces a greenish fluorescence (lambda(max) = 508 nm) when irradiated with long ultraviolet light. The fluorescence is due to the presence of a chromophore consisting of an imidazolone ring, formed by a post-translational modification of the tripeptide -Ser(65)-Tyr(66)-Gly(67)-. GFP has been used extensively as a reporter protein for monitoring gene expression in eukaryotic and prokaryotic cells, but relatively little is known about the chemical mechanism by which fluorescence is produced, To obtain a better understanding of this problem, we studied a peptide fragment of GFP bearing the chromophore and a synthetic model compound of the chromophore, The results indicate that the GFP chromophore consists of an imidazolone ring structure and that the light emitter is the singlet excited state of the phenolate anion of the chromophore, Further, the light emission is highly dependent on the microenvironment around the chromophore and that inhibition of isomerization of the exo-methylene double bond of the chromophore accounts for its efficient light emission.