Kim Y M, Jeong S H, Yamazaki I, Piette L H, Han S, Hong S J
Department of Chemistry and Biochemistry, Utah State University, Logan 84322-0300.
Free Radic Res. 1995 Jan;22(1):11-21. doi: 10.3109/10715769509147524.
The 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) spin adduct of myoglobin (Mb) or hemoglobin (Hb) was formed when metmyoglobin (MetMb) or methemoglobin (MetHb) reacted with H2O2 in the presence of DMPO, and both decayed with half-life of a few minutes. The DMPO spin adduct of Mb decayed with biphasic kinetics with k1 = 0.645 min-1 and k2 = 0.012 min-1, indicating that the spin adduct consisted of two kinetically heterogeneous species, stable and unstable ones. The DPMO spin adduct of Hb, however, was homogeneous. Decay of both spin adducts was accelerated in the presence of tyrosine, tryptophan or cysteine, but not phenylalanine, methionine or histidine. The decay obeyed the first order kinetics at varying concentrations of the spin adducts. The decay was accelerated by denaturation and proteolysis of protein moiety. The decay rate was not affected by the extra addition of MetMb or MetHb to each spin adduct. The decay rate of the spin adduct of Mb was increased by hematin in the presence of H2O2 and decreased by catalase. Decay of stable spin adduct of Mb, however, was not significantly changed under any experimental conditions used. These results led us to conclude that instability of the DMPO-spin adducts of Mb and Hb is due to intramolecular redox reactions between the spin adducts and amino acid residues and/or products of the reaction between heme and H2O2.
当高铁肌红蛋白（MetMb）或高铁血红蛋白（MetHb）在5,5 - 二甲基 - 1 - 吡咯啉 - N - 氧化物（DMPO）存在下与H₂O₂反应时，会形成肌红蛋白（Mb）或血红蛋白（Hb）的DMPO自旋加合物，且二者均在几分钟的半衰期内衰变。Mb的DMPO自旋加合物以双相动力学衰变，k₁ = 0.645 min⁻¹，k₂ = 0.012 min⁻¹，这表明自旋加合物由两种动力学性质不同的物种组成，即稳定的和不稳定的物种。然而，Hb的DPMO自旋加合物是均匀的。在酪氨酸、色氨酸或半胱氨酸存在下，两种自旋加合物的衰变加速，但在苯丙氨酸、甲硫氨酸或组氨酸存在下则不然。在自旋加合物浓度变化时，衰变遵循一级动力学。蛋白质部分的变性和蛋白水解会加速衰变。向每个自旋加合物中额外添加MetMb或MetHb不会影响衰变率。在H₂O₂存在下，血红素会增加Mb自旋加合物的衰变率，而过氧化氢酶则会降低其衰变率。然而，在任何实验条件下，Mb稳定自旋加合物的衰变均无显著变化。这些结果使我们得出结论，Mb和Hb的DMPO - 自旋加合物的不稳定性是由于自旋加合物与氨基酸残基之间的分子内氧化还原反应和/或血红素与H₂O₂反应的产物所致。
