The amounts of NG-methylarginine derivatives in myelin basic protein (MBP) purified from dysmyelinating mutant and different stages of normal myelinating mouse brains have been studied by using h.p.l.c. with a highly sensitive post-column o-phthaldialdehyde derivative-formation method. All three naturally occurring derivatives (NG-monomethylarginine (MeArg), NGN′G-dimethylarginine [Me2(sym)Arg] and NGNG-dimethylarginine [Me2(asym)Arg]) were found in MBP; however, their relative concentrations varied significantly with the age of the animal. The amounts of MeArg and Me2(sym)Arg in MBP increased as a function of the age of the brain, whereas that of Me2(asym)Arg decreased. MBP from early-myelinating mouse brain was shown to contain a high proportion of Me2(asym)Arg, which was hardly detectable in older brain MBP. This derivative, Me2(asym)Arg, was also absent from MBP embedded in the most compact multilamellar myelin, but was present in MBP in the least compact myelin (P3B). Comparing the extent of total methylation in vivo (sum of all three arginine derivatives), MBP extracted from less-compact myelin (P3A and P3B) showed a level approx. 40% higher than that from compact myelin. MBPs isolated from dysmyelinating mutant mouse brains, such as jimpy (jp/y) and quaking (qk/qk), contained a much higher level of Me2(asym)Arg relative to the other two methyl derivatives and also in comparison with those levels in the mother brain MBP. SDS/PAGE analysis of MBPs extracted from the mutant (both jp/y and qk/qk) as well as young normal (6-13 days old) mouse brains indicated the presence of a high-molecular-mass isoform of MBP (about 32 kDa), but this isoform was not found in adult brains. These results therefore indicate that structural integrity of myelin membrane in which MBP is embedded appears to play a pivotal role in determining the extent and the kind of Me2Arg formation in MBP at the post-translational level.
Studies on NG-methylarginine derivatives in myelin basic protein from developing and mutant mouse brain