NMR investigation of the charge isomers of bovine myelin basic protein.

Myelin basic protein (MBP) isolated from bovine white matter is obtained as a mixture of molecules which can be separated by cation-exchange chromatography at basic pH into three or more charge isomers. The three principal charge isomers of the microheterogeneous myelin basic protein have been isolated, and compared individually by high-resolution H NMR spectroscopy (360 and 400 MHz). In addition to confirming sources of MBP charge microheterogeneity such as fractional deamidation of Gln and loss of C-terminal Arg, NMR difference and spin-echo spectra further suggested (i) the presence of significant oxidation of (both) MBP Met residues to methionine sulfoxide; (ii) the three charge isomers contain equal ratios and absolute contents of mono- and dimethylated Arg; and (iii) the most-cationic isomer is deficient in its content of a putative extra Ala residue vs the other two isomers. Spectral analysis suggested that each MBP charge isomer is itself not a unique molecule, but more likely a mixture of molecules of equal net charge which are modified at any of the indicated functional side chains throughout the 169-residue protein. The results are discussed with respect to the possible consequences of MBP microheterogeneity to protein conformation and function.