title: Deimination of myelin basic protein. 2. Effect of methylation of MBP on its deimination by peptidylarginine deiminase.
abstract: Deimination of myelin basic protein (MBP) has been implicated in the chemical pathogenesis of multiple sclerosis (MS). Degradation of bovine MBP by cathepsin D, a myelin-associated protease, was increased when 6 arginyl residues were deiminated and became very rapid when all 18 arginyl residues were deiminated. Since MBP contains a number of modifications, including methylation, phosphorylation, etc., we studied the effect of methylation, an irreversible modification, to determine how this modification affected deimination. Methylation of Arg 106 in bovine MBP (Arg 107 in human), a naturally occurring modification of MBP, has been shown to affect the deimination of arginyl residues in the present study. Since fractionation of MBP into unmethylated, monomethylated, and dimethylated species cannot be done readily on a preparative scale, mass spectrometry with the Q-TOF instrument resolved these species readily since each differed from the other by 14 atomic mass units (amu). Examination of five different hMBP samples, two from normal brain and three from MS brain, revealed that increased deimination of arginyl residues correlated with a decreased methylation of Arg 107 (human sequence). To study this process in vitro, bovine MBP (bMBP) was used. Component 1 (C-1) is the most cationic of the MBP "charge isomers" and the most unmodified, in which all arginyl residues are intact. It was deiminated to various extents with purified bovine brain peptidylarginine deiminase, generating a number of species containing 0-13.7 mol of citrulline/mol of bMBP. Mass spectrometry of each of these species permitted us to determine the influence of methylation of Arg 106 (bovine sequence) on deimination by this enzyme. We found that bMBP with unmethylated arginine was deiminated at a rate of 0.081 mol of citrulline/min, with monomethylarginine, 0.068 mol of citrulline/min, and with dimethylarginine, 0.036 mol of citrulline/min. We suggest that the methylated arginyl residue becomes sequestered in the hydrophobic beta-sheet structure and disrupts the three-dimensional structure of the protein so that other arginyl residues are less accessible to peptidylarginine deiminase.
authors.affiliation: Department of Structural Biology & Biochemistry, The Hospital for Sick Children, Toronto, Ontario, Canada M5G 1X8.
authors.forename: L B
authors.initials: LB
authors.lastname: Pritzker
authors.forename: S
authors.initials: S
authors.lastname: Joshi
authors.forename: G
authors.initials: G
authors.lastname: Harauz
authors.forename: M A
authors.initials: MA
authors.lastname: Moscarello
mesh_terms: D000818:Animals; D001120:Arginine / Q000378:metabolism; D001921:Brain / Q000201:enzymology; D002402:Cathepsin D / Q000378:metabolism; D002417:Cattle; D002956:Citrulline / Q000378:metabolism; D006801:Humans; D006867:Hydrolases / Q000378:metabolism*; D007700:Kinetics; D013058:Mass Spectrometry; D008745:Methylation; D009103:Multiple Sclerosis / Q000201:enzymology; D004676:Myelin Basic Protein / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases
chemical_list: D004676:Myelin Basic Protein; D010446:Peptide Fragments; D002956:Citrulline; D001120:Arginine; D006867:Hydrolases; D002402:Cathepsin D; C000629235:PADI4 protein, human; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases