Elucidation of the Catalytic Function and the Inhibition mechanism of Phospholipase A_2

磷脂酶A_2的催化功能和抑制机制的阐明

基本信息

批准号：
05671853
负责人：
IKEDA Kiyoshi
金额：
$ 1.28万
依托单位：
Osaka university of Pharmaceutical Sciences
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05671853/
关键词：
Phospholipase A_2 Enzyme Inhibitor Substrate Analog Manoalide Catalytic Function Calcium Ion

项目摘要

1. Effects of Ca^<2+> and pH on the kinetic parameter for the hydrolysis of monodispersed 1,2-dihexanoy1-sn-glycero-3-phosphorylcholine, catalyzed by Group I and II phospholipases A_2 (PLA_2s) , were studied by the pH-stat assay method in the absence or presence of carbonic amidetype, oxazolidinone-type, and sulfonic amide-type substrate analogs. The Ca^<2+> dependency and participation of the catalytic group His 48 in the binding of genuine substrate to both types of PLA_2s were found to be very similar to those of the oxazolidinone-type substrate analog, but differed greatly from the carbonic amide-type and sulfonic amide-type substrate analogs. This finding suggests that the binding mode of oxazolidinone-type substrate analog is very similar to that of the genuine substrate.2. Chemical modification and inactivation of Group I and II PLA_2s were investigated by the use of a manoalide (MLD) -analog. It was found that Lys-56 of bovine pancreatic PLA_2s was modified by MLD-analog and that this modification was responsible for enzyme inactivation. It was indicated that the inactivation of pancreatic and N.naja atra PLA_2s originated from the modification of Lys residues at the interfacial recognition site, and that the inactivation of P.australis, T.flavoviridis and V.russelli russelli PLA_2s arose from the modification of Lys residues at the catalytic site, interfacial recognition site and regions outside both sits. The inactivation of A.halys blomhoffii PLA_2s was assumed to be due to the modification of Lys residues outside the two sites described above. We thought that the modification of the Lys residues outside the above two sites give rise to conformational changes leading to inactivation

1. Effects of Ca^<2+> and pH on the kinetic parameter for the hydrolysis of monodispersed 1,2-dihexanoy1-sn-glycero-3-phosphorylcholine, catalyzed by Group I and II phospholipases A_2 (PLA_2s) , were studied by the pH-stat assay method in the absence or presence of carbonic amidetype, oxazolidinone-type,和硫代酰胺类式底物类似物。发现催化群的Ca^<2+>依赖性和参与他的48在真正的底物与两种类型的PLA_2的结合中的结合非常相似，与黄选择素蛋白型型底物类似物的结合非常相似，但与碳纤维酰胺型和硫磺酰胺型酰胺型和硫磺酰胺 - 酰胺 - 酰胺 - 酰胺 - 酰胺 - 酰胺型型替代型相似。这一发现表明，黄唑啉酮型底物类似物的结合模式与真正的底物的结合模式非常相似。2。通过使用Manoalide（MLD）-Analog研究了I组和II组的化学修饰和灭活。发现牛胰腺PLA_2S的LYS-56通过MLD-Analog修饰，并且这种修饰是酶失活的原因。 It was indicated that the inactivation of pancreatic and N.naja atra PLA_2s originated from the modification of Lys residues at the interfacial recognition site, and that the inactivation of P.australis, T.flavoviridis and V.russelli russelli PLA_2s arose from the modification of Lys residues at the catalytic site, interfacial recognition site and regions outside both坐着。假定a.halys blomhoffii pla_2s的失活是由于上述两个位点以外的LYS残基的修饰。我们认为，在上述两个位点以外的LYS残基的修饰会导致构象变化，从而导致失活