CAJ | 학술논문

木聚糖是潜在的重要可再生能源,如何提高其降解效率已成为近年来的研究热点.β-木糖苷酶是木聚糖降解过程中的关键酶之一,按其水解机制可分为保留型与反转型酶.目前虽然对于这两种催化机制的研究不断深入,但很少有工作从溶液环境的角度出发探究它们的差异.本文采用分子动力学模拟方法,对4个典型的β-木糖苷酶进行了显式溶剂模拟研究,详细分析了酶的催化氨基酸间的距离和质子供体氨基酸pKa值的动态变化.结果显示,反转型酶催化氨基酸间的距离约为0.8-1.0 nm,大于保留型的0.5-0.6 nm,与先前对糖苷酶晶体结构的统计分析结果一致.令人意外的是,保留型酶的质子供体通过与其附近组氨酸的相互作用,其pKa在两个不同的高、低值之间交替变换,使保留型酶的双取代反应得以发生；而反转型酶的质子供体则由附近的天冬氨酸调节,其pKa稳定在某个较高值,这可能有利于其在反应pH值下获得水溶液中的氢离子,进行反转型酶特有的单取代反应.因此,本工作加深了人们对β-木糖苷酶保留型与反转型水解机制的认识,并为后续酶的理性改造与高效利用提供具有指导价值的结构与机理信息.
목취당시잠재적중요가재생능원,여하제고기강해효솔이성위근년래적연구열점.β-목당감매시목취당강해과정중적관건매지일,안기수해궤제가분위보류형여반전형매.목전수연대우저량충최화궤제적연구불단심입,단흔소유공작종용액배경적각도출발탐구타문적차이.본문채용분자동역학모의방법,대4개전형적β-목당감매진행료현식용제모의연구,상세분석료매적최화안기산간적거리화질자공체안기산pKa치적동태변화.결과현시,반전형매최화안기산간적거리약위0.8-1.0 nm,대우보류형적0.5-0.6 nm,여선전대당감매정체결구적통계분석결과일치.령인의외적시,보류형매적질자공체통과여기부근조안산적상호작용,기pKa재량개불동적고、저치지간교체변환,사보류형매적쌍취대반응득이발생；이반전형매적질자공체칙유부근적천동안산조절,기pKa은정재모개교고치,저가능유리우기재반응pH치하획득수용액중적경리자,진행반전형매특유적단취대반응.인차,본공작가심료인문대β-목당감매보류형여반전형수해궤제적인식,병위후속매적이성개조여고효이용제공구유지도개치적결구여궤리신식.
Xylans are important as potential renewable energy sources. In recent years, there has therefore been interest in improving their degradation efficiencies.β-Xylosidases are key enzymes for xylan degradation;these enzymes are classified, based on their hydrolysis mechanisms, as retaining or inverting enzymes. Although much research has been devoted to understanding retaining and inverting mechanisms, little is known about their differences in solution. We used molecular dynamics (MD) simulations with explicit solvent representation to study the dynamic behaviors of the active-sites of four typicalβ-xylosidases by analyzing the distances between two catalytic amino acids and the pKa values of proton-donor amino acids. The results show that the distance between the catalytic amino acids with inverting enzymes is about 0.8-1.0 nm, which is greater than that for retaining enzymes, i.e., 0.5-0.6 nm. This is consistent with previous results based on the crystal structures of glycosidases. We found that the pKa of the retaining proton donors are modulated by interactions with neighboring amino acids, enabling switching between low and high values. Such a pKa switch is needed for the double-displacement mechanism of retaining enzymes. In contrast, inverting proton donors, modulated by interactions with neighboring glutamic acids, have only high pKa values. This may be important in proton capture from the solvent by donors, and may facilitate the single-displacement mechanism of inverting enzymes. This study provides new insights into the hydrolysis mechanisms ofβ-xylosidases, and wil therefore be useful in improving the efficiency and applications ofβ-xylosidases.