鲢鱼和青鱼内源性转谷氨酰胺酶纯化及酶学性质比较  

Purification and comparison of enzymatic properties of endogenous transglutaminase between silver carp and black carp

作　　者：易林 安玥琦[1,2] 刘茹 胡杨[1,2] 熊善柏[1,2] YI Lin;AN Yueqi;LIU Ru;HU Yang;XIONG Shanba(College of Food Science and Technology,Huazhong Agricultural University,Wuhan,Hubei 430070,China;National R&D Branch Center for Conventional Freshwater Fish Processing〔Wuhan〕,Huazhong Agricultural University,Wuhan,Hubei 430070,China)

机构地区：[1]华中农业大学食品科学技术学院,湖北武汉430070 [2]华中农业大学国家大宗淡水鱼加工技术研发分中心〔武汉〕,湖北武汉430070

出　　处：《食品与机械》2023年第10期4-12,共9页Food and Machinery

基　　金：财政部和农业农村部:国家现代产业技术体系(编号:CARS-45-28)。

摘　　要：目的:探究鲢鱼、青鱼内源性转谷氨酰胺酶(transglutaminase,TGase)的酶学性质差异。方法:采用80%(NH_(4))_(2)SO_(4)盐析、Q-Sepharose FF和Sephacryl S-200 HR层析法从鲢鱼、青鱼肌肉中分离纯化出鲢鱼TGase(STG)、青鱼TGase(BTG),并对酶的相对分子质量、肽段序列、二级结构、适宜反应条件、热失活动力学等指标进行测定。结果:纯化后的STG和BTG的比酶活分别为14.34,12.67 U/mg,二者具有相近的相对分子质量。二者的肽段序列存在一定差异,其二级结构均以β-折叠为主,但STG的β-折叠含量略高于BTG的。STG和BTG的适宜反应温度均为50℃,适宜反应pH分别为8.0,7.5,完全激活两种TGase活性所需的Ca^(2+)浓度均为1 mmol/L,DTT可使两种TGase的酶活性增强,而PMSF、NH_(4)Cl、NEM、EDTA、Cu^(2+)、Ba^(2+)、Zn^(2+)、Mg^(2+)则会抑制其酶活。当温度为37~50℃时,热处理对STG和BTG的钝化均符合一级指数衰减动力学,二者的动力学参数E a值相近。结论:STG和BTG的一级结构及二级结构差异明显,但仍具有相似的适宜反应条件和热失活动力学特征。Objective:This study aimed to investigate the differences in enzymatic properties of endogenous transglutaminase(TGase)in silver carp and black carp.Methods:STG and BTG were purified from the muscle of silver carp and black carp,respectively,by 80%ammonium sulfate precipitation,Q-Sepharose FF,and Sephacryl S-200 HR chromatographies.Two enzymes were analyzed for relative molecular weights,peptide sequences,secondary structures,optimal reaction conditions,and thermal inactivation kinetics.Results:The purified STG and BTG showed similar relative molecular weights,of which the enzyme activities were 14.34 U/mg and 12.67 U/mg,respectively.Both enzymes showed differences in peptide sequences.The secondary structures of them were mainly theβ-fold,though the content ofβ-fold in STG was slightly higher than that of BTG.The optimal temperatures for STG and BTG were both 50℃,and the optimal pH values were 8.0 and 7.5,respectively.The enzymes required Ca^(2+)up to 1 mmol/L for full activation.The activities of STG and BTG were enhanced by DTT,whereas PMSF,NH_(4)Cl,NEM,EDTA,Cu^(2+),Ba^(2+),Zn^(2+),and Mg^(2+)showed inhibitory effects.When the temperature was 37~50℃,the passivations of STG and BTG by thermal treatment conformed to the first-order exponential decay kinetics with similar values of E a.Conclusion:The primary and secondary structures of STG and BTG exhibited obvious differences,yet they still exhibited similar properties in terms of optimal reaction conditions and thermal inactivation kinetics.

关 键 词：淡水鱼 转谷氨酰胺酶 分离纯化 结构特性 适宜反应条件 失活动力学 

分 类 号：TS254.1[轻工技术与工程—水产品加工及贮藏工程]