人肌和兔肌肌酸激酶在低pH条件下再折叠的研究

利用蛋白质内源荧光和远紫外ＣＤ光谱研究了在低ｐＨ条件下人肌肌酸激酶和兔肌肌酸激酶的构象状态。结果表明，在低离子强度下，随着酸的加入人肌和兔肌肌酸激酶去折叠，至ｐＨ２．０附近几乎都达到充分去折叠。它们的色氨酸荧光发射峰位红移至３５０ｎｍ，这表明了内埋的色氨酸残基已经完全暴露到极性溶剂之中，它们的远紫外ＣＤ光谱表明二级结构也遭破坏，但是仍保留有相当部分的二级结构。而且在高离子强度低ｐＨ条件下由Ｇｏｔｏ等人首先发现的中间体构象状态（融球结构状态）在我们的实验中也被观察到了。它具有与天然酶类似的二级结构。色氨酸荧光发射光谱表明与天然酶类似，它的最大发射峰位也为３３５ｎｍ表明了蛋白质已经完全折叠，然而其荧光强度远低于天然酶，表明这种结构状态具有较大运动性而导致荧光的动态淬灭。上述结果支持了Ｇｏｔｏ等人的发现，说明了融球中间体结构可能是蛋白质折叠过程需经历的一个中间态。

A STUDY ON THE FOLDING OF CREATINE KINASES FROM HUMAN AND RABBIT MUSCLE AT LOW pH

The confonnational stabal of cleatjne the from human and rabbit muscle at acidic pHun examined by the intrinsic proch fluoascence and far--ultlaviolet cellular dichIDism(far--UV CD). The adults we obtained show that at low iohic singtin, by adding HCl, tonative ha and rabbit creatine kinals ash proteins tended to adopt almost an fillly tmfolded confonnational state in the vicely of PH 2.0. Their tryptophan fluorescenceemission had a lhawh at the indicating a complete exposure of the nyptophan asiduesto the solvent. Thed far--UV CD spectla shoal partly disonled secondary str uperthg, butthe se dena Proteins retained soak secondary strucks. However, at high ionic strength andlow PH an in inte confonnational state (molten-globule state) which was fnSt discoverby Goto et al un also olived in our pat experiment . This intointe had a secondary thestructute similar to that of the nahve enzyme. The tryptophan fluorescence spectla were similar,with a mhanum at 335nm to that of the native on, indicating that the protein is almostrefolded complmp. Hoal, the intenSity of fluorescence elnjssion was much lower than thato f native enzyme, indicating that this state is more mobile and adulting in dynamicfluo quenching. The present adults support Goto's discovery. It is suggeStal thatmolten-globule state opt be an nounal folding lute-iate state of proteins.