两种微生物源谷氨酰胺转氨酶与底物特异性识别的全原子动力学模拟研究  

New insight into the substrate recognition mechanism of two microbe derived transglutaminases by an all-atoms dynamics simulation analysis

作　　者：吴楠 张永峰 季宝成 WU Nan;ZHANG Yong-Feng;JI Bao-Cheng(College of Food and Bioengineering,Zhengzhou University of Light Industry,Zhengzhou 450001,China)

机构地区：[1]郑州轻工业大学食品与生物工程学院,郑州450001

出　　处：《原子与分子物理学报》2023年第5期53-61,共9页Journal of Atomic and Molecular Physics

基　　金：国家自然科学基金(31901767);郑州轻工业大学博士科研启动基金(2018BSJJ020)。

摘　　要：微生物源谷氨酰胺转氨酶(Transglutaminase,TGase)能够催化蛋白间氨酰基转移促进蛋白交联,已经广泛应用于食品、轻工等多种领域.然而目前微生物源TGase与底物间基于长程作用力的分子特异性识别机制仍不清楚,理性改造困难.本研究以源自高茂源链霉菌(MTG)和源自枯草芽孢杆菌(BTG)的两种TGase为研究对象,使用分子对接技术构建酶与底物的结合复合体并使用分子动力学模拟解析两种TGase与底物识别的分子机制.结果显示两种TGase在底物识别过程高度类似,均由催化腔外周loop区域氨基酸残基通过范德华及静电作用固定底物整体,而后由位于催化腔底部催化三联体内Asp(MTG)或Glu(BTG)通过羧基侧链稳定反应发生局部空间构象并拉近底物Gln内γ-酰胺基与核心催化残基Cys间距,以便后续反应过程中TGase-底物中间体的生成.上述结果阐明了TGase与底物活性基团的分子识别模式,提出了两种TGase内作用于底物识别的氨基酸残基,为以后TGase的理性改造研究提供了理论基础.Transglutaminase(TGase)is a class of enzymes capable of catalyzing an amide transfer-based protein cross-linking,and thus is widely used in many fields.However,the substrate recognition mechanism of TGase remains unclear,hindering its rational molecular redesign.Here,we focus on two well-known TGases derived from Streptomyces mobara(MTG)and Bacillus subtilis(BTG),respectively,and docked them with a substrate CBZ-Gln-Gly.All-atom dynamics simulations in water were then performed using GROMACS software to characterize the structural and kinetic properties of the TGase-substrate complex.As a result,we found that both MTG and BTG employed similar substrate recognition mechanisms:the substrate initially bound to the external catalytic cavity because residues on this region donated strong electrostatic and Van der Walls interactions with it,while the Gln-γ-carboxyamide group of substrate was thereby orienting towards the catalytic triad at the bottom of the cavity.Asp(MTG)or Glu(BTG)in the catalytic triad attracted and grouped Cys(belonging to the catalytic triad and is a key residue for catalysis)and the Gln-γ-carboxyamide group of the substrate by forming hydrogen bond,therefore providing the short enough distance for covalent bond forming between TGase and substrate.Our findings demonstrated the binding properties between TGase and substrate,and identified critical residues respectively on MTG/BTG that contribute to substrate binding.

关 键 词：微生物源谷氨酰胺转氨酶 分子对接 分子动力学模拟 MM/PBSA aNCI分析 

分 类 号：Q615[生物学—生物物理学]