PDF(Pubmed)
Abstract：
BACKGROUND: Choline oxidase, a flavoprotein, is an enzyme that catalyzes the reaction which converts choline into glycine betaine. Choline oxidase started its journey way back in 1933. However, the impact of the high temperature on its structure has not been explored despite the long history and availability of its crystal structure. Both choline oxidase and its product, glycine betaine, have enormous applications spanning across multiple industries. Understanding how the 3D structure of the enzyme will change with the temperature change can open new ways to make it more stable and useful for industry.
METHODS: This research paper presents the in-silico study and analysis of the structural changes of A. globiformis choline oxidase at temperatures from 25 °C to 60 °C. A step-wise process is depicted in Fig. 1.
RESULTS: Multiple sequence alignment (MSA) of 11 choline oxidase sequences from different bacteria vs Arthrobacter globiformis choline oxidase showed that active site residues are highly conserved. The available crystal structure of A. globiformis choline oxidase with cofactor Flavin Adenine Dinucleotide (FAD) in the dimeric state (PDB ID: 4MJW)1 was considered for molecular dynamics simulations. A simulated annealing option was used to gradually increase the temperature of the system from 25 °C to 60 °C. Analysis of the conserved residues, as well as residues involved in Flavin Adenine Dinucleotide (FAD) binding, substrate binding, substate gating, and dimer formationwas done. At high temperatures, the formation of the inter-chain salt bridge between Arg50 and Glu63 was a significant observation near the active site of choline oxidase.
CONCLUSIONS: Molecular dynamics studies suggest that an increase in temperature has a significant impact on the extended Flavin Adenine Dinucleotide (FAD) binding region. These changes interfere with the entry of substrate to the active site of the enzyme and make the enzyme inactive.
METHODS: This research paper presents the in-silico study and analysis of the structural changes of A. globiformis choline oxidase at temperatures from 25 °C to 60 °C. A step-wise process is depicted in Fig. 1.
RESULTS: Multiple sequence alignment (MSA) of 11 choline oxidase sequences from different bacteria vs Arthrobacter globiformis choline oxidase showed that active site residues are highly conserved. The available crystal structure of A. globiformis choline oxidase with cofactor Flavin Adenine Dinucleotide (FAD) in the dimeric state (PDB ID: 4MJW)1 was considered for molecular dynamics simulations. A simulated annealing option was used to gradually increase the temperature of the system from 25 °C to 60 °C. Analysis of the conserved residues, as well as residues involved in Flavin Adenine Dinucleotide (FAD) binding, substrate binding, substate gating, and dimer formationwas done. At high temperatures, the formation of the inter-chain salt bridge between Arg50 and Glu63 was a significant observation near the active site of choline oxidase.
CONCLUSIONS: Molecular dynamics studies suggest that an increase in temperature has a significant impact on the extended Flavin Adenine Dinucleotide (FAD) binding region. These changes interfere with the entry of substrate to the active site of the enzyme and make the enzyme inactive.
摘要：
背景：胆碱氧化酶，一种黄素蛋白，是一种催化将胆碱转化为甘氨酸甜菜碱的反应的酶。胆碱氧化酶始于1933年。然而,尽管其晶体结构具有悠久的历史和可用性，但尚未探索高温对其结构的影响。胆碱氧化酶及其产物，甘氨酸甜菜碱,拥有跨越多个行业的巨大应用。了解酶的3D结构如何随温度变化而变化，可以开辟新的途径，使其更稳定，更有用。
方法：本研究论文介绍了在25°C至60°C的温度下，球形A胆碱氧化酶结构变化的计算机研究和分析。在图1中描绘了一个逐步的过程。1.
结果：来自不同细菌的11个胆碱氧化酶序列与球形节杆菌胆碱氧化酶的多序列比对（MSA）表明，活性位点残基高度保守。对于分子动力学模拟,考虑了具有处于二聚体状态的辅因子黄素腺嘌呤二核苷酸(FAD)1(PDBID:4MJW)1的球形曲霉胆碱氧化酶的可用晶体结构。使用模拟退火选项将系统的温度从25°C逐渐增加到60°C。分析保守的残基，以及涉及黄素腺嘌呤二核苷酸（FAD）结合的残基，底物结合，子状态门控，二聚体形成完成。在高温下,Arg50和Glu63之间链间盐桥的形成是胆碱氧化酶活性位点附近的重要观察结果。
结论：分子动力学研究表明，温度升高对延伸的黄素腺嘌呤二核苷酸（FAD）结合区具有显着影响。这些变化干扰底物进入酶的活性位点并使酶失活。
方法：本研究论文介绍了在25°C至60°C的温度下，球形A胆碱氧化酶结构变化的计算机研究和分析。在图1中描绘了一个逐步的过程。1.
结果：来自不同细菌的11个胆碱氧化酶序列与球形节杆菌胆碱氧化酶的多序列比对（MSA）表明，活性位点残基高度保守。对于分子动力学模拟,考虑了具有处于二聚体状态的辅因子黄素腺嘌呤二核苷酸(FAD)1(PDBID:4MJW)1的球形曲霉胆碱氧化酶的可用晶体结构。使用模拟退火选项将系统的温度从25°C逐渐增加到60°C。分析保守的残基，以及涉及黄素腺嘌呤二核苷酸（FAD）结合的残基，底物结合，子状态门控，二聚体形成完成。在高温下,Arg50和Glu63之间链间盐桥的形成是胆碱氧化酶活性位点附近的重要观察结果。
结论：分子动力学研究表明，温度升高对延伸的黄素腺嘌呤二核苷酸（FAD）结合区具有显着影响。这些变化干扰底物进入酶的活性位点并使酶失活。
