Department of Biological Chemistry, University of Michigan Medical Center, Ann Arbor, MI 48109-5606, USA.
Proc Natl Acad Sci U S A. 2009 Dec 22;106(51):21567-72. doi: 10.1073/pnas.0908106106. Epub 2009 Dec 2.
The mechanism by which docking fidelity is achieved for the multitude of cofactor-dependent enzymes is poorly understood. In this study, we demonstrate that delivery of coenzyme B(12) or 5'-deoxyadenosylcobalamin by adenosyltransferase to methylmalonyl-CoA mutase is gated by a small G protein, MeaB. While the GTP-binding energy is needed for the editing function; that is, to discriminate between active and inactive cofactor forms, the chemical energy of GTP hydrolysis is required for gating cofactor transfer. The G protein chaperone also exerts its editing function during turnover by using the binding energy of GTP to elicit release of inactive cofactor that is occasionally formed during the catalytic cycle of MCM. The physiological relevance of this mechanism is demonstrated by a patient mutation in methylmalonyl-CoA mutase that does not impair the activity of this enzyme per se but corrupts both the fidelity of the cofactor-loading process and the ejection of inactive cofactor that forms occasionally during catalysis. Consequently, cofactor in the incorrect oxidation state gains access to the mutase active site and is not released if generated during catalysis, leading, respectively, to assembly and accumulation of inactive enzyme and resulting in methylmalonic aciduria.
对于众多依赖辅因子的酶来说，其对接保真度的实现机制尚未完全阐明。在本研究中，我们证明了腺苷转移酶将辅酶 B(12)或 5'-脱氧腺苷钴胺素递送至甲基丙二酰辅酶 A 变位酶是由一种小 G 蛋白 MeaB 控制的。虽然 GTP 结合能对于编辑功能（即区分活性和非活性辅因子形式）是必需的，但 GTP 水解的化学能对于门控辅因子转移是必需的。G 蛋白伴侣在周转过程中也发挥其编辑功能，利用 GTP 的结合能来引发偶尔在 MCM 催化循环中形成的非活性辅因子的释放。这种机制的生理相关性通过甲基丙二酰辅酶 A 变位酶的患者突变得到了证明，该突变本身并不损害该酶的活性，但会破坏辅因子加载过程的保真度以及在催化过程中偶尔形成的非活性辅因子的释放。因此，处于错误氧化态的辅因子进入变位酶的活性位点，如果在催化过程中生成则不会被释放，从而导致活性酶的组装和积累，导致甲基丙二酸尿症。
