Abstract：
Amino acid epimerization, a process of converting L-amino acids to D-amino acids, will lead to modification in the protein structure and, subsequently, its biological function. This modification causes no change in protein m/z and may be overlooked during protein analysis. Aspartic Acid Epimerization (AAE) is faster than other amino acids and could be accelerated by free radicals and peroxides. In this work, a novel and site-specific HPLC method using a chiral stationary phase for determining the AAE in the active site model peptide (AP) of Peroxiredoxin 2 has been developed and validated. The developed method showed good linearity (1 - 200 μg/mL) and recoveries of the limit of quantification (LOQ), low, medium, and high concentrations were between 85% and 115%. The Kinetics of AAE in AP were studied using the developed method, and the results showed that when ascorbic acid and Cu2+ coexisted, the AP epimerized rapidly. The AAE extent increased with time and was positively correlated with hydrogen peroxide generation.
摘要：
氨基酸差向异构化，将L-氨基酸转化为D-氨基酸的过程,会导致蛋白质结构的改变，随后,其生物学功能。这种修饰不会引起蛋白质m/z的变化，并且在蛋白质分析过程中可能会被忽略。天冬氨酸差向异构化（AAE）比其他氨基酸更快，并且可以被自由基和过氧化物加速。在这项工作中,已开发并验证了一种新颖的位点特异性HPLC方法，该方法使用手性固定相确定过氧化物酶2的活性位点模型肽（AP）中的AAE。该方法具有良好的线性(1-200μg/mL)和定量限(LOQ),低,中等,高浓度在85%到115%之间。使用开发的方法研究了AP中AAE的动力学，结果表明，当抗坏血酸和Cu2+共存时，AP的empimerized迅速。AAE程度随时间增加，与过氧化氢的产生呈正相关。
