Stimulation of bovine brain phospholipase C activity by myelin basic protein requires arginyl residues in peptide linkage.
We reported previously a highly purified phosphatidylinositol-specific phospholipase C (PI-PLC) from bovine brain and from human myelin which was stimulated by myelin basic protein. In this paper we report that the stimulation of the PI-PLC activity by myelin basic protein (MBP) requires arginine residues in peptide linkage. MBP and poly-L-arginine were able to stimulate the PI-PLC activity by 250% while other basic poly amino acids were unable to stimulate the PI-PLC activity. Neither free arginine nor benzoyl-arginine ethylester was able to stimulate the activity of the enzyme. These results suggested a requirement for the guanidino group of arginine and arginine in peptidyl linkage. The arginyl residues of MBP were modified chemically with 1,2-cyclohexanedione, or enzymatically by cholera toxin which ADP-ribosylated arginyl groups, or by peptidylarginine deiminase which converted the guanidino group of arginine to the ureido group of citrulline. ADP-ribosylation did not affect the stimulation while the 1,2-cyclohexanedione modified MBP and the peptidylarginine deiminase-treated MBP showed a reduced ability to stimulate the PI-PLC activity which correlated with the number of arginyl residues modified. Sequence analysis of the peptidylarginine deiminase-treated MBP established that specific arginyl residues had been converted to citrulline to a greater extent than others. When 70% of Arg 25 and Arg31 were converted to citrulline little stimulatory activity remained, whereas the conversion of 100% of Arg 170 did not affect the ability of C1 to stimulate the enzyme. A role for "active" arginine in this MBP peptide is suggested by our data.
Tompkins
T A
TA
Department of Biochemistry, Hospital for Sick Children, Toronto, Canada.
Moscarello
M A
MA
D000818:Animals; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology*; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D002956:Citrulline / Q000378:metabolism; D006867:Hydrolases / Q000378:metabolism; D004676:Myelin Basic Protein / Q000494:pharmacology*; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology*; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000187:drug effects / Q000378:metabolism*; D011107:Polylysine / Q000494:pharmacology; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases; D017421:Sequence Analysis; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000378:metabolism
D004676:Myelin Basic Protein; D010455:Peptides; C008973:polyornithine; D011107:Polylysine; C015462:polyarginine; D002956:Citrulline; D001120:Arginine; D006867:Hydrolases; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D014357:Trypsin; C000629235:PADI4 protein, human; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases; D043265:Phosphatidylinositol Diacylglycerol-Lyase