Abstract：
Limited alliinase resources cause difficulties in the biosynthesis of thiosulfinates (e.g., allicin), restricting their applications in the agricultural and food industries. To effectively biosynthesize thiosulfinates, this study aimed to excavate bacterial alliinase resources and elucidate their catalytic properties. Two bacterial cystathionine β-lyases (MetCs) possessing high alliinase activity (>60 U mg -1) toward L-(-)-alliin were identified from Allium sativum rhizosphere isolates. Metagenomic exploration revealed that cystathionine β-lyase from Bacillus cereus (BcPatB) possessed high activity toward both L-(±)-alliin and L-(+)-alliin (208.6 and 225.1 U mg -1), respectively. Although these enzymes all preferred l-cysteine S-conjugate sulfoxides as substrates, BcPatB had a closer phylogenetic relationship with Allium alliinases and shared several similar features with A. sativum alliinase. Interestingly, the Trp30Ile31Ala32Asp33 Met34 motif in a cuspate loop of BcPatB, especially sites 31 and 32 at the top of the motif, was modeled to locate near the sulfoxide of L-(+)-alliin and is important for substrate stereospecificity. Moreover, the stereoselectivity and activity of mutants I31V and A32G were higher toward L-(+)-alliin than those of mutant I31L/D33E toward L-(-)-alliin. Using bacterial alliinases and chemically synthesized substrates, we obtained thiosulfinates with high antimicrobial and antinematode activities that could provide insights into the protection of crops and food.
摘要：
有限的蒜氨酸酶资源导致硫代亚磺酸酯的生物合成困难(例如，大蒜素)，限制了它们在农业和食品工业中的应用。为了有效地生物合成硫代亚磺酸酯，本研究旨在挖掘细菌蒜氨酸酶资源，阐明其催化特性。从大蒜根际分离株中鉴定出两种对L-（-）-alliin具有高alliinase活性（>60Umg-1）的细菌cystathionineβ-裂解酶（MetCs）。宏基因组学研究表明，蜡样芽孢杆菌（BcPatB）对L-（±）-蒜氨酸和L-（）-蒜氨酸（208.6和225.1Umg-1）均具有较高的活性，分别。尽管这些酶都优选l-半胱氨酸S-缀合亚砜作为底物，BcPatB与Alliumalliinase具有更紧密的系统发育关系，并且与A.sativumalliinase具有相似的特征。有趣的是,Trp30Ile31Ala32Asp33Met34基序在BcPatB的尖点环中，尤其是主题顶部的31和32，被建模为位于L-（）-alliin的亚砜附近，对于底物立体特异性很重要。此外，突变体I31V和A32G对L-（）-alliin的立体选择性和活性高于突变体I31L/D33E对L-（-）-alliin的立体选择性和活性。使用细菌蒜氨酸酶和化学合成的底物，我们获得了具有高抗微生物和抗线虫活性的硫代亚磺酸盐，可以为作物和食物的保护提供见解。
