Department of Life Sciences, University of Bath, Claverton Down, Bath BA2 7AY, UK.
Biomolecules. 2024 Mar 2;14(3):299. doi: 10.3390/biom14030299.
α-Methylacyl-CoA racemase in (MCR) has an essential role in fatty acid metabolism and cholesterol utilization, contributing to the bacterium's survival and persistence. Understanding the enzymatic activity and structural features of MCR provides insights into its physiological and pathological significance and potential as a therapeutic target. Here, we report high-resolution crystal structures for wild-type MCR in a new crystal form (at 1.65 Å resolution) and for three active-site mutants, H126A, D156A and E241A, at 2.45, 1.64 and 1.85 Å resolutions, respectively. Our analysis of the new wild-type structure revealed a similar dimeric arrangement of MCR molecules to that previously reported and details of the catalytic site. The determination of the structures of these H126A, D156A and E241A mutants, along with their detailed kinetic analysis, has now allowed for a rigorous assessment of their catalytic properties. No significant change outside the enzymatic active site was observed in the three mutants, establishing that the diminution of catalytic activity is mainly attributable to disruption of the catalytic apparatus involving key hydrogen bonding and water-mediated interactions. The wild-type structure, together with detailed mutational and biochemical data, provide a basis for understanding the catalytic properties of this enzyme, which is important for the design of future anti-tuberculosis drug molecules.
α-甲基酰基辅酶 A 消旋酶（MCR）在脂肪酸代谢和胆固醇利用中发挥着重要作用，有助于细菌的生存和持续存在。了解 MCR 的酶学活性和结构特征，有助于深入了解其生理和病理意义及其作为治疗靶点的潜力。在这里，我们报道了野生型 MCR 在一种新晶体形式下的高分辨率晶体结构（分辨率为 1.65Å）以及三个活性位点突变体 H126A、D156A 和 E241A 的晶体结构，分辨率分别为 2.45Å、1.64Å 和 1.85Å。我们对新的野生型结构的分析揭示了 MCR 分子的类似二聚体排列，以及催化位点的细节。这些 H126A、D156A 和 E241A 突变体的结构测定及其详细的动力学分析，现在可以对其催化特性进行严格评估。在三个突变体中，在酶活性位点之外没有观察到明显的变化，这表明催化活性的降低主要归因于涉及关键氢键和水介导相互作用的催化装置的破坏。野生型结构以及详细的突变和生化数据为理解该酶的催化特性提供了基础，这对于设计未来的抗结核药物分子至关重要。
