Department of Biochemistry, Faculty of Science, P.J. Šafárik University, Moyzesova 11, 041 54 Košice, Slovakia.
Institute of Microbiology - BioCeV, Vídeňská 1083, 142 20 Prague 4, Czech Republic.
Int J Biol Macromol. 2021 Mar 31;174:413-423. doi: 10.1016/j.ijbiomac.2021.01.189. Epub 2021 Jan 30.
Cytochrome c (cyt c), in addition to its function as an electron shuttle in respiratory chain, is able to perform as a pseudo-peroxidase with a critical role during apoptosis. Incubation of cyt c with an excess of hydrogen peroxide leads to a suicide inactivation of the protein, which is accompanied by heme destruction and covalent modification of numerous amino acid residues. Although steady-state reactions of cyt c with an excess of hydrogen peroxide represent non-physiological conditions, they might be used for analysis of the first-modified amino acid in in vivo. Here, we observed oxidation of tyrosine residues 67 and 74 and heme as the first modifications found upon incubation with hydrogen peroxide. The positions of the oxidized tyrosines suggest a possible migration pathway of hydrogen peroxide-induced radicals from the site of heme localization to the protein surface. Analysis of a size of folded fraction of cyt c upon limited incubation with hydrogen peroxide indicates that the early oxidation of amino acids triggers an accelerated destruction of cyt c. Position of channels from molecular dynamics simulation structures of cyt c points to a location of amino acid residues exposed to reactive oxidants that are thus more prone to covalent modification.
细胞色素 c（cyt c）除了在呼吸链中作为电子穿梭体发挥作用外，还能够作为一种拟过氧化物酶发挥作用，在细胞凋亡过程中具有关键作用。cyt c 与过量的过氧化氢孵育会导致蛋白质的自杀失活，伴随着血红素的破坏和许多氨基酸残基的共价修饰。尽管 cyt c 与过量的过氧化氢的稳态反应代表非生理条件，但它们可能用于分析体内第一个被修饰的氨基酸。在这里，我们观察到过氧化氢孵育后第一个被修饰的是酪氨酸残基 67 和 74 的氧化和血红素。氧化酪氨酸的位置表明，过氧化氢诱导的自由基可能从血红素定位的位置迁移到蛋白质表面的一种可能途径。对 cyt c 进行有限的过氧化氢孵育后的折叠部分大小的分析表明，早期的氨基酸氧化会触发 cyt c 的加速破坏。分子动力学模拟结构中通道的位置指向了暴露于反应性氧化剂的氨基酸残基的位置，因此更容易发生共价修饰。
