通过有限蛋白酶解评估金属离子诱导的大肠杆菌碱性磷酸酶构象变化。
Metal-ion induced conformational changes in alkaline phosphatase from E. coli assessed by limited proteolysis.
作者信息
Bucević-Popović V, Pavela-Vrancic M, Dieckmann R
机构信息
Department of Chemistry, Faculty of Natural Sciences, Mathematics and Education, University of Split, N. Tesle 12, 21000 Split, Croatia.
出版信息
Biochimie. 2004 Jun;86(6):403-9. doi: 10.1016/j.biochi.2004.05.001.
Alkaline phosphatase (AP) displays significant structural changes during metal-ion binding, supporting cooperative interactions between the subunits of the dimeric enzyme. Here, we present data on the dynamic properties of AP from E. coli, and characterize the structural changes that accompany variations in metal-ion content, combining limited proteolysis and MALDI-TOF mass spectrometry. Limited proteolysis revealed an internal cleavage site at Arg-293, reflecting a position of conformational flexibility supporting subunit communication essential for catalysis. A specific shielding of a region distant from the metal-binding site has been demonstrated, implying transmission of conformational changes, induced by metal-ion binding to the adjacent subunit, across the subunit interface.
碱性磷酸酶（AP）在金属离子结合过程中表现出显著的结构变化，这支持了二聚体酶亚基之间的协同相互作用。在此，我们展示了来自大肠杆菌的AP的动力学特性数据，并结合有限蛋白酶解和基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS），对伴随金属离子含量变化的结构变化进行了表征。有限蛋白酶解揭示了在精氨酸-293处的一个内部切割位点，这反映了一个构象灵活性的位置，该位置支持对催化至关重要的亚基通讯。已证明远离金属结合位点的一个区域存在特异性屏蔽，这意味着由金属离子与相邻亚基结合诱导的构象变化会通过亚基界面进行传递。
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