Hille R, Anderson R F
Department of Medical Biochemistry, Ohio State University, Columbus 43210.
J Biol Chem. 1991 Mar 25;266(9):5608-15.
Electron transfer within milk xanthine oxidase has been examined by the technique of pulse radiolysis. Radiolytically generated N-methylnicotinamide radical or 5-deazalumiflavin radical has been used to rapidly and selectively introduce reducing equivalents into the enzyme so that subsequent equilibration among the four redox-active centers of the enzyme (a molybdenum center, two iron-sulfur centers, and FAD) could be monitored spectrophotometrically. Experiments have been performed at pH 6 and 8.5, and a comprehensive scheme describing electron equilibration within the enzyme at both pH values has been developed. All rate constants ascribed to equilibration between specific pairs of centers in the enzyme are found to be rapid relative to enzyme turnover under the same conditions. Electron equilibration between the molybdenum center and one of the iron-sulfur centers of the enzyme (tentatively assigned Fe/S I) is particularly rapid, with a pH-independent first-order rate constant of approximately 8.5 x 10(3) s-1. The results unambiguously demonstrate the role of the iron-sulfur centers of xanthine oxidase in mediating electron transfer between the molybdenum and flavin centers of the enzyme.
已通过脉冲辐解技术研究了乳黄嘌呤氧化酶内的电子转移。辐射产生的N-甲基烟酰胺自由基或5-脱氮黄素自由基已被用于快速且选择性地将还原当量引入该酶中，以便随后能够通过分光光度法监测该酶的四个氧化还原活性中心（一个钼中心、两个铁硫中心和黄素腺嘌呤二核苷酸）之间的平衡。实验在pH 6和8.5条件下进行，并且已经制定了一个描述该酶在这两个pH值下电子平衡的综合方案。发现在相同条件下，相对于酶的周转而言，归因于该酶中特定中心对之间平衡的所有速率常数都很快。该酶的钼中心与其中一个铁硫中心（暂定为Fe/S I）之间的电子平衡特别快，其pH无关的一级速率常数约为8.5×10³ s⁻¹。结果明确证明了黄嘌呤氧化酶的铁硫中心在介导该酶的钼中心和黄素中心之间的电子转移中的作用。
