Protein conformational gating of enzymatic activity in xanthine oxidoreductase.
Protein conformational gating of enzymatic activity in xanthine oxidoreductase.
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DOI：
10.1021/ja207173p
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发表时间：
2012-01-18
影响因子：
15
通讯作者：
Pai EF
中科院分区：
文献类型：
--
作者：
Ishikita H;Eger BT;Okamoto K;Nishino T;Pai EF
在哺乳动物中，黄嘌呤氧化还原酶可以作为黄嘌呤脱氢酶（XDH）和黄嘌呤氧化酶（XO）存在。这两种酶具有共同的氧化还原活性辅因子，其形成由黄素辅因子终止的电子转移（ET）途径。尽管具有相同的蛋白质一级结构，但XDH和XO之间的黄素半醌/氢醌对的氧化还原电位差（Esq/hq）为~170 mV，这是一个显著的差异。前者非常喜欢NAD+作为最终底物，通过黄素从铁硫簇FeS-II，而后者只接受分子氧。然而，在XDH（无NAD+）中，电子供体FeS-II的氧化还原电位比受体黄素的氧化还原电位高180 mV，产生能量上的上坡ET。基于XDH、XO、NAD+-和NADH-复合的XDH的新的1.65、2.3、1.9和2.2 μ m分辨率晶体结构，计算Esq/hq以更好地理解酶如何将ET从FeS-II活化为黄素。XDH和XO之间的Esq/hq差异的大部分来源于在靠近黄素结合位点的位置423-433处的环中的构象变化，导致半醌状态的稳定性的差异。没有观察到响应于XDH处的NAD+结合的大的构象变化。相反，NAD+环的正电荷、Asp 429的去质子化和NAD+分子对黄素本体表面的加帽都有助于改变NAD+与XDH结合后的Esq/hq。
In mammals, xanthine oxidoreductase can exist as xanthine dehydrogenase (XDH) and xanthine oxidase (XO). The two enzymes possess common redox active cofactors, which form an electron transfer (ET) pathway terminated by a flavin cofactor. In spite of identical protein primary structures, the redox potential difference between XDH and XO for the flavin semi-quinone/hydroquinone pair (Esq/hq) is ~170 mV, a striking difference. The former greatly prefers NAD+ as ultimate substrate for ET from the iron-sulfur cluster FeS-II via flavin while the latter only accepts dioxygen. In XDH (without NAD+), however, the redox potential of the electron donor FeS-II is 180 mV higher than that for the acceptor flavin, yielding an energetically uphill ET. Based on new 1.65, 2.3, 1.9 and 2.2 Å resolution crystal structures for XDH, XO, the NAD+- and NADH- complexed XDH, Esq/hq were calculated to better understand how the enzyme activates an ET from FeS-II to flavin. The majority of the Esq/hq difference between XDH and XO originates from a conformational change in the loop at positions 423-433 near the flavin binding site, causing the differences in stability of the semiquinone state. There was no large conformational change observed in response to NAD+ binding at XDH. Instead, the positive charge of NAD+ ring, deprotonation of Asp429 and the capping of the bulk surface of the flavin by the NAD+ molecule all contribute to alter Esq/hq upon NAD+ binding to XDH.