*College of Medicine, Burnett School of Biomedical Sciences, 6900 Lake Nona Boulevard, Orlando, FL 32827, U.S.A.
Biochem J. 2013 Nov 15;456(1):129-37. doi: 10.1042/BJ20130981.
The dihaem enzyme MauG catalyses a six-electron oxidation required for post-translational modification of preMADH (precursor of methylamine dehydrogenase) to complete the biosynthesis of its TTQ (tryptophan tryptophylquinone) cofactor. Trp93 of MauG is positioned midway between its two haems, and in close proximity to a Ca2+ that is critical for MauG function. Mutation of Trp93 to tyrosine caused loss of bound Ca2+ and changes in spectral features similar to those observed after removal of Ca2+ from WT (wild-type) MauG. However, whereas Ca2+-depleted WT MauG is inactive, W93Y MauG exhibited TTQ biosynthesis activity. The rate of TTQ biosynthesis from preMADH was much lower than that of WT MauG and exhibited highly unusual kinetic behaviour. The steady-state reaction exhibited a long lag phase, the duration of which was dependent on the concentration of preMADH. The accumulation of reaction intermediates, including a diradical species of preMADH and quinol MADH (methylamine dehydrogenase), was detected during this pre-steady-state phase. In contrast, steady-state oxidation of quinol MADH to TTQ, the final step of TTQ biosynthesis, exhibited no lag phase. A kinetic model is presented to explain the long pre-steady-state phase of the reaction of W93Y MauG, and the role of this conserved tryptophan residue in MauG and related dihaem enzymes is discussed.
双血红素酶 MauG 催化六电子氧化反应，该反应是前 MADH（亚甲基胺脱氢酶的前体）翻译后修饰所必需的，以完成其 TTQ（色氨酸色基醌）辅因子的生物合成。MauG 中的色氨酸 93 位于两个血红素之间，并且与 Ca2+ 接近，该 Ca2+ 对 MauG 的功能至关重要。色氨酸 93 突变为酪氨酸导致结合的 Ca2+丢失，并改变了光谱特征，类似于从 WT（野生型）MauG 中除去 Ca2+后观察到的特征。然而，尽管 Ca2+耗尽的 WT MauG 无活性，但 W93Y MauG 表现出 TTQ 生物合成活性。从前 MADH 生成 TTQ 的速率远低于 WT MauG，并且表现出非常异常的动力学行为。稳态反应表现出长的迟滞期，其持续时间取决于前 MADH 的浓度。在此预稳态阶段检测到反应中间体的积累，包括前 MADH 的二自由基物种和醌 MADH（亚甲基胺脱氢酶）。相比之下，稳态氧化醌 MADH 生成 TTQ，即 TTQ 生物合成的最后一步，没有迟滞期。提出了一个动力学模型来解释 W93Y MauG 反应的长预稳态阶段，并且讨论了该保守色氨酸残基在 MauG 和相关双血红素酶中的作用。
