Department of Biochemistry and Molecular Biology, University of Parma, 43100 Parma, Italy.
Biochemistry. 2008 Jun 24;47(25):6560-70. doi: 10.1021/bi7023749.
Methylamine can be used as the sole carbon source of certain methylotrophic bacteria. Methylamine dehydrogenase catalyzes the conversion of methylamine into formaldehyde and donates electrons to the electron transfer protein amicyanin. The crystal structure of the complex of methylamine dehydrogenase and amicyanin from Paracoccus versutus has been determined, and the rate of electron transfer from the tryptophan tryptophylquinone cofactor of methylamine dehydrogenase to the copper ion of amicyanin in solution has been determined. In the presence of monovalent ions, the rate of electron transfer from the methylamine-reduced TTQ is much higher than in their absence. In general, the kinetics are similar to those observed for the system from Paracoccus denitrificans. The complex in solution has been studied using nuclear magnetic resonance. Signals of perdeuterated, (15)N-enriched amicyanin bound to methylamine dehydrogenase are observed. Chemical shift perturbation analysis indicates that the dissociation rate constant is approximately 250 s(-1) and that amicyanin assumes a well-defined position in the complex in solution. The most affected residues are in the interface observed in the crystal structure, whereas smaller chemical shift changes extend to deep inside the protein. These perturbations can be correlated to small differences in the hydrogen bond network observed in the crystal structures of free and bound amicyanin. This study indicates that chemical shift changes can be used as reliable indicators of subtle structural changes even in a complex larger than 100 kDa.
甲胺可以作为某些甲基营养细菌的唯一碳源。甲胺脱氢酶催化甲胺转化为甲醛，并将电子供体传递给电子转移蛋白血蓝蛋白。已确定来自 Paracoccus versutus 的甲胺脱氢酶和血蓝蛋白复合物的晶体结构，并确定了溶液中甲胺脱氢酶的色氨酸色氨酸醌辅因子向血蓝蛋白铜离子的电子转移速率。在单价离子存在下，甲胺还原的 TTQ 的电子转移速率远高于不存在时。一般来说，动力学与从 Paracoccus denitrificans 观察到的系统相似。已使用核磁共振研究了溶液中的复合物。观察到结合甲胺脱氢酶的氘代、(15)N 丰度的血蓝蛋白的信号。化学位移扰动分析表明，离解速率常数约为 250 s(-1)，并且在溶液中的复合物中血蓝蛋白占据明确的位置。受影响最大的残基位于晶体结构中观察到的界面处，而较小的化学位移变化延伸到蛋白质内部深处。这些扰动可以与游离和结合的血蓝蛋白的晶体结构中观察到的氢键网络中的微小差异相关联。这项研究表明，化学位移变化可用作可靠的指标，即使在大于 100 kDa 的复杂体系中，也能检测到细微的结构变化。
