金黄色葡萄球菌透明质酸裂解酶HylS的异源表达与特性研究  

Heterologous Expression and Characterization of the Hyaluronic Acid Lyase HylS from Staphylococcus aureus

作　　者：岑潇龙 雷曦 马诗云 陈倩茹 黄遵锡[1,2,3,4] 周峻沛 张蕊[1,2,3,4] CEN Xiao-long;LEI Xi;MA Shi-yun;CHEN Qian-ru;HUANG Zun-xi;ZHOU Jun-pei;ZHANG Rui(College of Life Science,Yunnan Normal University,Kunming 650500;Engineering Research Center of Sustainable Development and Utilization of Biomass Energy,Ministry of Education,Kunming 650500;Key Laboratory of Yunnan for Biomass Energy and Biotechnology of Environment,Kunming 650500;Key Laboratory of Yunnan Provincial Education Department for Plateau Characteristic Food Enzymes,Kunming 650500)

机构地区：[1]云南师范大学生命科学学院,昆明650500 [2]生物能源持续开发利用教育部工程研究中心,昆明650500 [3]云南省生物质能与环境生物技术重点实验室,昆明650500 [4]云南省高校高原特色食品酶重点实验室,昆明650500

出　　处：《生物技术通报》2022年第1期157-167,共11页Biotechnology Bulletin

基　　金：国家重点研发计划(2017YFB0308401);国家自然科学基金项目(32060214);云南省基础研究计划重点项目(202001AS070022);云南省万人计划青年拔尖人才(YNWR-QNBJ-2018-383);云南师范大学联大青年学者培养计划(01200205020516036)。

摘　　要：透明质酸酶能够将透明质酸聚糖降解成具有抗氧化等生物活性的低分子量寡糖。微生物来源透明质酸酶具有酶学性质多样和易于重组表达等特点,是开发透明质酸酶的研究热点。通过基因组测序获得一个潜在的金黄色葡萄球菌来源透明质酸裂解酶基因hylS,将其进行了大肠杆菌BL21(DE3)异源重组表达,并对重组酶进行了酶学特性和酶解产物抗氧化性分析。纯化后的重组酶rHylS的最适pH和温度分别为5.0和45℃;专一性降解透明质酸,比活是(1.6×10^(5)±5.4)U/mg;降解透明质酸产生低分子量的不饱和寡糖,属于内切透明质酸裂解酶;酶解产物对ABTS、DPPH、超氧阴离子和羟自由基清除能力显著高于未酶解的高分子量透明质酸,且与浓度呈正相关。金黄色葡萄球菌来源的透明质酸裂解酶HylS酶学性质优良,可用于生产具有抗氧化性低分子量的不饱和透明质酸寡糖。Hyaluronidases degrade hyaluronic acid into low molecular weight oligosaccharides with biological activities such as antioxidant.Hyaluronidases from microorganisms have been a hot area of research in the development of hyaluronidase,because of their diverse enzymatic properties and easily recombinant expression.A potential hyaluronic acid(HA)lyase(hylS)gene from Staphylococcus aureus was identified by genome sequencing.It was successfully expressed by Escherichia coli BL21(DE3).The characteristics of the recombinant enzyme were analyzed,and the antioxidation of its enzymolysis products was determined.The optimal pH and temperature of the purified recombinant enzyme rHylS was 5.0 and 45℃,respectively.It showed specific degradation of HA,and specific activity towards the substrate was(1.6×10^(5)±5.4)U/mg.It was an endo-hyaluronate lyase that degraded HA to produce low molecular weight unsaturated oligosaccharides(HAP).The scavenging abilities of HAP on ABTS,DPPH,superoxide anions and hydroxyl radicals were significantly higher than that of high molecular weight HA without degradation,and there was a positive correlation with the concentration.All these excellent characters indicate that rHylS from S.aureus is of enzymatic property,and it may be used to produce unsaturated hyaluronic acid oligosaccharides of low molecular weight and antioxidant properties.

关 键 词：金黄色葡萄球菌 透明质酸 裂解酶 酶学性质 不饱和寡糖 抗氧化性 

分 类 号：Q78[生物学—分子生物学]