CAJ | 학술논문

采用非变性聚丙烯酰胺凝胶电泳、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳、高效凝胶排阻色谱以及激光光散射光谱研究了脲变性牛碳酸酐酶B的稀释复性过程及其集聚作用.在脲变性牛碳酸酐酶B的稀释复性过程中,当最终复性液中脲浓度大于2.0 mol/L时,牛碳酸酐酶B在复性液中以单分子和二分子集聚体形式存在;当最终复性液中脲浓度小于2.0 mol/L大于1.0 mol/L时,牛碳酸酐酶B在复性液中以单分子、二分子集聚体和少量多分子集聚体形式存在;而当最终复性液中脲浓度小于等于1.0 mol/L时,脲变性牛碳酸酐酶B复性时会形成均匀透明的上清和不透明的沉淀,牛碳酸酐酶B在上清和沉淀中达到动态解离平衡,且在两相中都以单分子、二分子集聚体和少量多分子集聚体形式存在.溶液中二分子和多分子牛碳酸酐酶B集聚体是通过牛碳酸酐酶B分子之间的疏水和静电相互作用力而形成的,当溶液中这些成分达到一定浓度并且溶液中脲的浓度小于某一个值时,它们之间会通过非共价形式形成沉淀.
채용비변성취병희선알응효전영、십이완기류산납-취병희선알응효전영、고효응효배조색보이급격광광산사광보연구료뇨변성우탄산항매B적희석복성과정급기집취작용.재뇨변성우탄산항매B적희석복성과정중,당최종복성액중뇨농도대우2.0 mol/L시,우탄산항매B재복성액중이단분자화이분자집취체형식존재;당최종복성액중뇨농도소우2.0 mol/L대우1.0 mol/L시,우탄산항매B재복성액중이단분자、이분자집취체화소량다분자집취체형식존재;이당최종복성액중뇨농도소우등우1.0 mol/L시,뇨변성우탄산항매B복성시회형성균균투명적상청화불투명적침정,우탄산항매B재상청화침정중체도동태해리평형,차재량상중도이단분자、이분자집취체화소량다분자집취체형식존재.용액중이분자화다분자우탄산항매B집취체시통과우탄산항매B분자지간적소수화정전상호작용력이형성적,당용액중저사성분체도일정농도병차용액중뇨적농도소우모일개치시,타문지간회통과비공개형식형성침정.
The dilution refolding and aggregation interaction of bovine carbonic anhydrase B induced by urea were studied by using non-denatured PAGE, SDS-PAGE, size-exclusion chromatography and laser light-scattering. When the final urea concentration in renaturation solutions was higher than 2.0 mol/L, bovine carbonic anhydrase B existed in the renaturation solutions in unimolecular form and bi-molecular aggregate; when the final urea concentration was in the range of 1.0-2.0 mol/L, they existed in a unimolecular form, bi-molecular aggregate and a small amount of multi-molecular aggregate; and when the final urea concentration was lower or equal to 1.0 mol/L, a transparent supernatant and an opaque precipitate were separately formed in the dilution refolding of urea-induced bovine carbonic anhydrase B, a dynamic dissociation equilibrium existed in the bovine carbonic anhydrase B included in the supernatant and that included in the precipitate and in both the supernatant and precipitate, and bovine carbonic anhydrase B existed in a unimolecular form, bi-molecular and a small amount of multi-molecular aggregate. The bi-molecular and multi-molecular aggregates were formed through the hydrophobic and electrostatic interactions between the unimolecular bovine carbonic anhydrase B, and only if the concentrations of these components in renaturation solution were higher than a certain value and the urea concentration in the final renaturation solution was lower than a certain value, an aggregate precipitate could be formed through the non-covalent interactions among these components.