力竭游泳诱导的大鼠睾丸金属结合蛋白的初步分离与鉴定  

Separation and identification of testicular metal-binding proteins induced by exhaustive swimming in rats

作　　者：张天欧[1,2] 温悦萌[1,2] 谢岚[1,2] 梁伟锋[3] 艾华[1,2] 

机构地区：[1]北京大学第三医院运动医学研究所,北京100191 [2]北京大学肥胖与代谢病研究中心营养运动与肥胖研究室,北京100191 [3]北京蛋白质组研究中心,北京102206

出　　处：《中国实验动物学报》2012年第3期1-6,91,共7页Acta Laboratorium Animalis Scientia Sinica

基　　金：国家自然科学基金(编号:39670185)

摘　　要：目的初步分离和鉴定力竭运动诱导的大鼠睾丸金属结合蛋白(testis metal-binding proteins,TMB-Ps)。方法 8只雄性SD大鼠一次性力竭游泳运动(8只对照)后6 h取睾丸和肝脏组织,用镉饱和法测定TMBPs和肝脏金属硫蛋白(metallothionein,MT)含量,用tricine-SDS-PAGE方法分离镉饱和法提取液的TMBPs组分和肝脏MT,用液相色谱-串联质谱法(LC-MS-MS)鉴定TMBPs分离条带。结果力竭运动组大鼠TMBPs水平(113.71±11.72)nmol Cd/g testis显著高于安静对照组(87.14±12.72)nmol Cd/g testis(P<0.01),肝脏MT表达量(64.70±14.89)μg/g也显著高于安静对照组(7.32±3.31)μg/g(P<0.001)。LC-MS-MS对tricine-SDS-PAGE分离的蛋白条带的鉴定结果为:TMBPs含有泛素、铜-锌-超氧化物歧化酶、酪蛋白样磷蛋白等蛋白质,另外还应包括MT。结论TMBPs由一组具有金属结合性、耐热性和诱导性的蛋白质所组成,主要有泛素、超氧化物歧化酶和酪蛋白样磷蛋白。镉饱和法并非测定金属硫蛋白的特异性方法。Objective To separate and identify the characteristics of testicular metal-binding proteins（TMBPs） induced by exhaustive swimming in rats.Methods Eight male SD rats were subjected to a bout of exhaustive swimming（8 for control） and the testes and liver were taken out 6 hours after the exercise.TMBPs and hepatic metallothionein（MT） were determined by cadmium-saturation assay.The extracts from the testes and liver by cadmium-saturation method were separated by tricine-SDS-PAGE and TMBPs protein bands were identified by LC-MS-MS.Results The results of cadmium-saturation assay showed that the TMBPs content was significantly increased in the exhaustive swimming rats [（113.71±11.72）nmol Cd/g testis] than that in the controls [（87.14±12.72）nmol Cd/g testis]（P〈0.01）,and the hepatic MT was（64.70±14.89）μg/g,significantly higher than the（7.32±3.31）μg/g in the controls（P〈0.001）.Ubiquitin,superoxide dismutase （Cu-Zn-SOD）,casein-like phosphoprotein（CLPH） were identified in TMBPs,and MT was also included.Conclusions TMBPs are mainly composed of ubiquitin,SOD and CLPH,with characteristics of metal binding,heat resistance and induction.The cadmium-saturation assay is not a specific method for MT determination.

关 键 词：睾丸金属结合蛋白 金属硫蛋白 蛋白质组学 镉饱和法 

分 类 号：Q95-33[生物学—动物学]