人肌肌酸激酶在SDS溶液中失活与构象变化的比较研究

应用紫外差吸收光谱、荧光发射光谱、ＣＤ先谱等监测手段，研究了ＳＤＳ溶液滴定人肌肌酸激酶时的构象与活力变化的关系。结果表明酶的活力丧失先于以紫外差吸收先谱、荧先发射谱和巯基暴露数目所监测到的构象变化。ＳＤＳ滴定时引起的酶的荧光发射光谱的变化在低滴定度阶段随着ＳＤＳ滴定量的增加，荧光强度下降，发射峰位红移，当ＳＤＳ浓度达到２．１ｍｍｏｌ／Ｌ时，荧光强度增大，继续增加ＳＤＳ滴定量，荧光强度又降低，发射峰位红移直至终态。紫外差吸收光谱随着ＳＤＳ溶液的加入，２８１ｎｍ．２８７ｎｍ和２９２ｎｍ的负差吸收峰增大。ＣＤ光谱结果表明在本实验所用的ＳＤＳ浓度范围内，ＳＤＳ对人肌肌酸激酶的二级结构几乎没有影响。上述结果支持了酶的活性部位构象柔性的观点。

CONFORMATIONAL AND ACTIVITY CHANGES OF HUMAN CREATINE KINASE DURING SDS DENATURATION

The conformational changes of human creatine kinase during titration by Sodium Dodecyl Sulfate(SDS)solution were studied by ulhaviolet difference spectra ,intrin sic protein fluorescence and Circular Dicchrosm (CD). The adults showed that changes of conformation measured in abothance of 287nm, the red shift of fluorelnhaon maximaand the nUmbers of expend SH groups were much fasfor than that of activity.Changes in the fluormce emission spectla of the during Sac titration occur in tWO stagals.decade of emission intenSity took place at low Sac concentration with with shill of ion maxim,when the concentration of Sin reached tO 2. Innnol/L, the emission intensity fllrther changes in fluoascence propend occur at a more higher Sin contion.The CD special showed that the secondary strUctWh of enzyme in our expeconcentlation of Sin had no obvious change. The present results supports the condition that the conformational state of the active site of is very fragile and easily disruted.