Evdokimov Artem G, Pokross Matthew E, Egorov Nikolay S, Zaraisky Andrey G, Yampolsky Ilya V, Merzlyak Ekaterina M, Shkoporov Andrey N, Sander Ian, Lukyanov Konstantin A, Chudakov Dmitriy M
X-ray crystallography, HCRC, Discovery, Procter&Gamble Pharmaceuticals, 8700 Mason-Montgomery Road, Mason, Ohio 45040, USA.
EMBO Rep. 2006 Oct;7(10):1006-12. doi: 10.1038/sj.embor.7400787. Epub 2006 Aug 25.
Since the cloning of Aequorea victoria green fluorescent protein (GFP) in 1992, a family of known GFP-like proteins has been growing rapidly. Today, it includes more than a hundred proteins with different spectral characteristics cloned from Cnidaria species. For some of these proteins, crystal structures have been solved, showing diversity in chromophore modifications and conformational states. However, we are still far from a complete understanding of the origin, functions and evolution of the GFP family. Novel proteins of the family were recently cloned from evolutionarily distant marine Copepoda species, phylum Arthropoda, demonstrating an extremely rapid generation of fluorescent signal. Here, we have generated a non-aggregating mutant of Copepoda fluorescent protein and solved its high-resolution crystal structure. It was found that the protein beta-barrel contains a pore, leading to the chromophore. Using site-directed mutagenesis, we showed that this feature is critical for the fast maturation of the chromophore.
自1992年维多利亚多管水母绿色荧光蛋白（GFP）被克隆以来，已知的类GFP蛋白家族一直在迅速壮大。如今，该家族包含了从刺胞动物物种中克隆出的一百多种具有不同光谱特征的蛋白。对于其中一些蛋白，其晶体结构已被解析，显示出在发色团修饰和构象状态方面的多样性。然而，我们对GFP家族的起源、功能和进化仍远未完全了解。该家族的新型蛋白最近从进化关系较远的海洋桡足类物种（节肢动物门）中克隆出来，显示出荧光信号的极快速产生。在此，我们生成了一种桡足类荧光蛋白的非聚集突变体，并解析了其高分辨率晶体结构。结果发现，该蛋白的β桶包含一个通向发色团的孔。通过定点诱变，我们表明这一特征对于发色团的快速成熟至关重要。
