光谱法研究变性剂诱导的猪胃蛋白酶和牛血清蛋白折叠中间态的分布和过渡

Spectrophotometic Investigation on the Stable Conformations of Porcine Pepsin and Bovine Serum Albumin Induced by Denaturation Solutions

利用紫外光谱和荧光光谱分别测定了变性剂诱导的猪胃蛋白酶和牛血清蛋白的残余活性,并根据它们的两个特征展开参数定量描述了猪胃蛋白酶和牛血清蛋白天然态、折叠中间态和完全展开态随变性液中变性剂浓度的分布和过渡。结果表明,在盐酸胍诱导的猪胃蛋白酶"三态"去折叠过程中,当盐酸胍浓度约为1.5mol/L时,折叠中间态的浓度达到最大,约占溶液中总猪胃蛋白酶的12%;在脲诱导的牛血清蛋白"三态"去折叠过程中,当脲浓度约为2.0mol/L时,折叠中间态的浓度达到最大,约占溶液中总牛血清蛋白的41%;而在盐酸胍诱导的牛血清蛋白"四态"去折叠过程中,当盐酸胍浓度约为0.4mol/L、1.2mol/L时,第一和第二折叠中间态浓度分别达到最大,约占溶液中总牛血清蛋白的20%和70%。

The residual activities of porcine pepsin and bovine serum albumin induced by denaturant were studied with the UV spectroscopy or fluorescence spectroscopy. By means of the characteristic unfolding parameters,the distribution and transition of stable conformatioas of porcine pepsin and bovine serum albumin under different denaturant concentrations were well described. The results shows that during the three-states-unfolding of porcine pepsin induced by guanidine hydrochloride, the maximum（12%） intermediate content is obtained when the concentration of guanidine hydrochloride is 1.5 mol/L; during the three-states-unfolding of bovine serum albumin induced by urea,the maximum（41%） intermediate content is obtained when the concentration of urea is 2. 0 mol/L; during the four-states-unfolding of bovine serum albumin induced by guanidine hydrochloride, when the concentration of guanidine hydrochloride is approximately 0.4 mol/L and 1.2 mol/L,the maximum intermediate I1 and I2 content is achieved, accounting for 20 % and 70 %, respectively.