title: Sphingomyelin modulates interfacial binding of Taiwan cobra phospholipase A2.
abstract: The goal of the present study is to elucidate the effect of sphingomyelin on interfacial binding of Taiwan cobra phospholipase A(2) (PLA(2)). Substitution of Asn-1 with Met caused a reduction in enzymatic activity and membrane-damaging activity of PLA(2) toward phospholipid vesicles, while sphingomyelin exerted an inhibitory effect on the biological activities of native and mutated PLA(2). Incorporation of sphingomyelin reduced membrane fluidity of phospholipid vesicles as evidenced by Laurdan fluorescence measurement. The results of self-quenching studies, binding of fluorescent probe, trinitrophenylation of Lys residues and fluorescence energy transfer between protein and lipid revealed that sphingomyelin altered differently membrane-bound mode of native and mutated PLA(2). Moreover, it was found that PLA(2) and N-terminally mutated PLA(2) adopted different conformation and geometrical arrangement on binding with membrane bilayer. Nevertheless, the binding affinity of PLA(2) and N-terminal mutant for phospholipid vesicles was not greatly affected by sphingomyelin. Together with the finding that mutation on N-terminus altered the gross conformation of PLA(2), our data indicate that sphingomyelin modulates the mode of membrane binding of PLA(2) at water/lipid interface, and suggest that the modulated effect of sphingomyelin depends on inherent structural elements of PLA(2).
authors.affiliation: Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan.
authors.forename: Yi-Ling
authors.initials: YL
authors.lastname: Chiou
authors.forename: Shinne-Ren
authors.initials: SR
authors.lastname: Lin
authors.forename: Long-Sen
authors.initials: LS
authors.lastname: Chang
mesh_terms: D000818:Animals; D020134:Catalytic Domain; D004546:Elapid Venoms / Q000201:enzymology*; D005454:Fluorescence Polarization; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D008560:Membrane Fluidity; D044367:Phase Transition; D054467:Phospholipases A2 / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D013109:Sphingomyelins / Q000737:chemistry*; D013624:Taiwan; D013696:Temperature
chemical_list: D004546:Elapid Venoms; D005456:Fluorescent Dyes; D013109:Sphingomyelins; D054467:Phospholipases A2