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.

Arginine Methylation of Yeast mRNA-binding Protein Npl3 Directly Affects Its Function, Nuclear Export, and Intranuclear Protein Interactions

“…Non-histone substrates of both type I and type II methyltransferases can also be targets for deimination (54,61), but in vitro data suggest that arginine dimethylation inhibits deimination (60,61). We did not detect citrulline residues within Npl3 by mass spectrometry and S. cerevisiae has no obvious PAD homolog.…”

Multiple Sclerosis

“…n, number of samples. ation had a protective effect on MBP (41). Therefore the apparent increase in methylation in the MS samples may reflect the greater resistance to proteolysis by methylation, imparting a longer half-life to the protein.…”

Extracellular Matrix Remodeling: The Common Denominator in Connective Tissue DiseasesPossibilities for Evaluation and Current Understanding of the Matrix as More Than a Passive Architecture, but a Key Player in Tissue Failure

“…Many of these modifications have been implicated in the antigenicity of the proteins, as outlined in Table 4 (modified from Cloos and Christgau 15 ). 296,311,[319][320][321][322]346,347,[376][377][378][379][380][381][382][383][384][385][386][387][388][389] These observations have sparked a growing interest in the role and assessment of PTMs in autoimmune diseases as well as other pathological conditions associated with aging. Whether the presence of PTMs is merely a secondary phenomenon accompanying the disease or a primary event in disease initiation remains to be resolved.…”