Abstract：
Spontaneous deamidation of amino acids is a physiologically important process, particularly for protein aging and diseases. Despite its widespread occurrence, the mechanism of glutamine deamidation particularly within proteins remains poorly understood. We have used a multiscale computational approach to investigate glutamine deamidation in the tripeptide Glycine-Glutamine-Glycine (Gly-Gln-Gly) and γS-Crystallin protein. Specifically, both the 5- and 6-membered water-assisted deamidation pathways in the tripeptide have been elucidated and compared. Both are found to occur in three stages: iminol formation, cyclization, and deamination. The rate-limiting step in each mechanism is nucleophilic attack of the backbone iminol nitrogen, formed in the first stage, at the glutamine\'s side-chain carbonyl carbon. For the 6- and 5-membered mechanisms, this occurs with a free energy cost of 136.4 and 179.5 kJ mol-1, respectively. Thus, overall, in the Gly-Gln-Gly tripeptide, the 6-membered pathway is preferred. Furthermore, the free energies for forming cyclic intermediates and products at selected Gln residues (based on experimentally reported % deamidation) in γS-Crystallin have been obtained. It is found that the 5-membered product complex is exergonic at -25.3 kJ mol-1, while the 6-membered product complex is calculated to be endergonic at 90.7 kJ mol-1. Thus, the deamidation pathway in folded and constrained proteins may not exclusively follow the 6-membered route. Molecular dynamics (MD) simulations of γS-Crystallin indicate that deamidation is more likely to occur when two or more water molecules are in the proximity of the glutamine residue. Consequently, significant conformational changes are found to accompany Gln120 deamidation in γS-Crystallin. This in turn can influence water availability at the other Gln residues considered and hence potentially their deamidation. Collectively, these results provide comprehensive insights into spontaneous water-assisted deamidation of glutamine residues in peptides and into the role and impact of Gln deamidation in proteins.
摘要：
氨基酸的自发脱酰胺是一个重要的生理过程，特别是蛋白质老化和疾病。尽管它普遍存在，谷氨酰胺脱酰胺的机制，特别是在蛋白质中仍然知之甚少。我们使用多尺度计算方法研究了三肽甘氨酸-谷氨酰胺-甘氨酸（Gly-Gln-Gly）和γS-Crystallin蛋白中的谷氨酰胺脱酰胺作用。具体来说,已经阐明并比较了三肽中的5元和6元水辅助脱酰胺化途径。发现两者都发生在三个阶段：亚氨基醇的形成，环化,和脱氨。每种机制中的限速步骤都是骨架亚氨基醇氮的亲核攻击，形成于第一阶段,在谷氨酰胺的侧链羰基碳。对于六元和五元机制，发生这种情况的自由能成本分别为136.4和179.5kJmol-1。因此，总的来说,在Gly-Gln-Gly三肽中，6元途径是优选的。此外，已获得在γS-Crystallin中选定的Gln残基上形成环状中间体和产物的自由能（基于实验报道的％脱酰胺作用）。发现5元产物络合物在-25.3kJmol-1时是exergonic，而6元产物络合物在90.7kJmol-1时是endergonic。因此，折叠和受限蛋白质的脱酰胺途径可能不完全遵循6元途径。γS-晶体蛋白的分子动力学（MD）模拟表明，当两个或多个水分子靠近谷氨酰胺残基时，脱酰胺作用更容易发生。因此,发现γS-Crystallin中的Gln120脱酰胺作用伴随着显着的构象变化。这反过来会影响所考虑的其他Gln残基处的水可用性,并因此潜在地影响它们的脱酰胺作用。总的来说，这些结果为肽中谷氨酰胺残基的自发水辅助脱酰胺化以及Gln脱酰胺化在蛋白质中的作用和影响提供了全面的见解。
