Department of Chemistry, University of Utah, 315 South 1400 East, Room 2020, Salt Lake City, Utah 84112, United States.
Radboud University, Institute for Molecules and Materials, FELIX Laboratory, Toernooiveld 7, 6525 ED Nijmegen, The Netherlands.
J Am Soc Mass Spectrom. 2021 Mar 3;32(3):786-805. doi: 10.1021/jasms.0c00468. Epub 2021 Feb 11.
Deamidation of asparaginyl (Asn) peptides is a spontaneous post-translational modification that plays a significant role in degenerative diseases and other biological processes under physiological conditions. In the gas phase, deamidation of protonated peptides is a major fragmentation channel upon activation by collision-induced dissociation. Here, we present a full description of the deamidation process from protonated asparagine-serine, [AsnSer+H], via infrared (IR) action spectroscopy and threshold collision-induced dissociation (TCID) experiments in combination with theoretical calculations. The IR results demonstrate that deamidation proceeds via bifurcating reaction pathways leading to furanone- and succinimide-type product ion structures, with a population analysis indicating the latter product dominates. Theory demonstrates that nucleophilic attack of the peptidyl amide oxygen onto the Asn side chain leads to furanone formation, whereas nucleophilic attack by the peptidyl amide nitrogen onto the Asn side-chain carbonyl carbon leads to the formation of the succinimide product structure. TCID experiments find that furanone formation has a threshold energy of 145 ± 12 kJ/mol and succinimide formation occurs with a threshold energy of 131 ± 12 kJ/mol, consistent with theoretical energies and with the spectroscopic results indicating that succinimide dominates. The results provide information regarding the inductive and steric effects of the Ser side chain on the deamidation process. The other major channel observed in the TCID experiments of [AsnSer+H] is dehydration, where a threshold energy of 104 ± 10 kJ/mol is determined. A complete IR and theoretical analysis of this pathway is also provided. As for deamidation, a bifurcating pathway is found with both dominant oxazoline and minor diketopiperazine products identified. Here, the Ser side chain is directly involved in both pathways.
天冬酰胺（Asn）肽的脱酰胺作用是一种自发的翻译后修饰，在生理条件下，它在退行性疾病和其他生物过程中起着重要作用。在气相中，质子化肽的脱酰胺作用是通过碰撞诱导解离（CID）激活的主要碎裂通道。在这里，我们通过红外（IR）作用光谱和阈能碰撞诱导解离（TCID）实验以及理论计算，对质子化天冬酰胺-丝氨酸[AsnSer+H]的脱酰胺过程进行了全面描述。IR 结果表明，脱酰胺作用通过分叉反应途径进行，导致呋喃酮和琥珀酰亚胺型产物离子结构，通过种群分析表明后者产物占主导地位。理论表明，肽酰胺氧对天冬酰胺侧链的亲核攻击导致呋喃酮形成，而肽酰胺氮对天冬酰胺侧链羰基碳的亲核攻击导致琥珀酰亚胺产物结构的形成。TCID 实验发现，呋喃酮的形成具有 145 ± 12 kJ/mol 的阈能，而琥珀酰亚胺的形成发生在 131 ± 12 kJ/mol 的阈能，与理论能量一致，与光谱结果表明琥珀酰亚胺占主导地位。结果提供了关于丝氨酸侧链对脱酰胺过程的诱导和空间位阻效应的信息。在[AsnSer+H]的 TCID 实验中观察到的另一个主要通道是脱水，确定其阈能为 104 ± 10 kJ/mol。还提供了对该途径的完整 IR 和理论分析。与脱酰胺作用一样，发现了一条分叉途径，确定了主要的恶唑啉和次要的二酮哌嗪产物。在这里，丝氨酸侧链直接参与了这两种途径。
