Chemical modification and inactivation of bovine pancreatic, porcine pancreatic, Naja naja atra and Pseudechis australis phospholipases A2 (PLA2s), belonging to Group I, and of Trimeresurus flavoviridis, Vipera russelli russelli and Agkistrodon halys blomhoffii PLA2s, belonging to Group II, were investigated by the use of a manoalide (MLD)-analogue, 1-(2,5-dihydro-hydroxy-5-oxo-3-furanyl)-8,12-dimethyl-4-formyl-3,7, 11-tridecatrienol. At appropriate time intervals, residual PLA2 activities towards monodispersed, anionic mixed micellar and non-ionic mixed micellar substrates were measured. We tested the protective effect of micellar n-dodecylphosphocholine (n-C12PC) on enzyme inactivation. Inactivation of pancreatic PLA2s (Group I) was only observed towards anionic mixed micellar substrates. This inactivation was completely prevented by the presence of micellar n-C12PC. From a fragmentation study of modified bovine pancreatic PLA2 using lysyl endopeptidase, we speculated that Lys-56 of this enzyme was modified by MLD-analogue and that this modification was responsible for enzyme inactivation. Inactivation of non-pancreatic PLA2s was observed towards all types of substrate, except that no significant inactivation of N. naja atra PLA2 (Group I) towards monodispersed substrate was noted. Micellar n-C12PC protected N. naja atra PLA2 (Group I) completely from inactivation by MLD-analogue, but had lesser protective effects on P. australis PLA2 (Group I), T. flavoviridis and V. russelli russelli PLA2s (Group II). However, no significant protection of A. halys blomhoffii PLA2s (Group II) activity was observed. These results indicate that the inactivation of pancreatic and N. naja atra PLA2s originates from the modification of Lys residues at the interfacial recognition site, and that inactivation of P. australis, T. flavoviridis and V. russelli PLA2s arises from the modification of Lys residues at the catalytic site, interfacial recognition site and regions outside both sites. The inactivation of A. halys blomhoffii PLA2 was assumed to be due to the modification of Lys residues outside the two sites described above.
Chemical modification and inactivation of phospholipases A2 by a manoalide analogue
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S Fujii, Y Tahara, M Toyomoto, S Hada, H Nishimura, S Inoue, K Ikeda, Y Inagaki, S Katsumura, Y Samejima, T Omori-Satoh, C Takasaki, K Hayashi; Chemical modification and inactivation of phospholipases A2 by a manoalide analogue. Biochem J 15 May 1995; 308 (1): 297–304. doi: https://doi.org/10.1042/bj3080297
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