芽孢杆菌中的酒石酸脱氢酶:通过动力学、结构和分子对接分析解密独特的催化多样性

IF 1.9 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY
Manali Chandnani, Disha Patel, Twinkle Patel, Aditi Buch
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引用次数: 0

摘要

分歧进化的酒石酸脱氢酶(TDH)在单个活性位点上表现出多种催化活性;来自普氏拟杆菌(P. putida)的酶(pTDH)在结构上和生物化学上都有很好的表征。由于芽孢杆菌分离物中存在与 TDH 相关的有氧代谢 L-酒石酸盐的能力,而且 ycsA 编码的蛋白质序列与 pTDH 的相似性有限,因此芽孢杆菌 TDH 可能是一种具有催化多样性和进化意义的令人感兴趣的酶。本研究通过动力学、结构和分子对接分析,探讨了枯草芽孢杆菌 168(168bTDH)和地衣芽孢杆菌 DSM-13(429bTDH)的 TDH 的底物相互作用。从大肠杆菌 BL21(DE3)细胞不溶性馏分中纯化的异源表达 bTDHs 在正向反应中能显著催化作为底物的 L-酒石酸盐和中酒石酸盐。与 pTDH 不同的是,bTDHs 在以二羟富马酸为底物的反向反应中的催化能力比 pTDH 高出约 4 倍。分子对接预测的结合能进一步证实了它们的相对底物特异性,同时揭示了活性位点上参与蛋白质-配体相互作用的主要残基。动力学分析和同源建模通过拉马钱德兰图分析进行了验证,预测 bTDH 具有二聚体性质。总之,这些结果凸显了系统发育较晚的 bTDHs 的独特催化潜力,为介导高效对映选择性酶促生物转化提供了一个重要的蛋白质工程目标。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Tartrate Dehydrogenase in Bacillus Species: Deciphering Unique Catalytic Diversity Through Kinetic, Structural and Molecular Docking Analysis

Tartrate Dehydrogenase in Bacillus Species: Deciphering Unique Catalytic Diversity Through Kinetic, Structural and Molecular Docking Analysis

Divergently evolved Tartrate dehydrogenase (TDH) exhibits multiple catalytic activities at a single active site; the enzyme from P. putida (pTDH) being structurally and biochemically well-characterized. Occurrence of TDH-associated ability to aerobically metabolize L-tartrate in Bacillus isolates and limited resemblance of ycsA-encoded protein sequences with pTDH rendered Bacillus TDH as an intriguing enzyme with possible catalytic diversity as well as evolutionary significance. The present study explores substrate interactions of TDHs from B. subtilis 168 (168bTDH) and B. licheniformis DSM-13 (429bTDH) through kinetic, structural and molecular docking-based analysis. Heterologously expressed bTDHs, purified from insoluble fractions of E. coli BL21(DE3) cells, could significantly catalyze L-tartrate and meso-tartrate as substrates in forward reaction. Unlike pTDH, bTDHs distinctly and more efficiently catalyzed the reverse reaction using dihydroxyfumarate substrate following sigmoidal kinetics; the ability being ~ 4 fold higher in 168bTDH. Their binding energies predicted from molecular docking, further substantiated the relative substrate specificities, while revealing major residues involved in protein-ligand interactions at active site. The kinetic analysis and homology modelling validated using Ramachandran Plot analysis predicted a dimeric nature for bTDH. Collectively, the results highlight unique catalytic potential of phylogenetically recent bTDHs, offering an important protein engineering target to mediate efficient enantioselective enzymatic biotransformations.

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来源期刊
The Protein Journal
The Protein Journal 生物-生化与分子生物学
CiteScore
5.20
自引率
0.00%
发文量
57
审稿时长
12 months
期刊介绍: The Protein Journal (formerly the Journal of Protein Chemistry) publishes original research work on all aspects of proteins and peptides. These include studies concerned with covalent or three-dimensional structure determination (X-ray, NMR, cryoEM, EPR/ESR, optical methods, etc.), computational aspects of protein structure and function, protein folding and misfolding, assembly, genetics, evolution, proteomics, molecular biology, protein engineering, protein nanotechnology, protein purification and analysis and peptide synthesis, as well as the elucidation and interpretation of the molecular bases of biological activities of proteins and peptides. We accept original research papers, reviews, mini-reviews, hypotheses, opinion papers, and letters to the editor.
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