Magnetic resonance study of glycophorin A-containing 13C-enriched methionines

Methionine-81 and/or -8 of the transmembrane sialoglycoprotein, glycophorin A, have been specifically alkylated with 13CH3I to produce the sulfonium ion derivatives [S-[13C]methylmethionine-8]glycophorin A and [S-[13C]methylmethionine-8 and -81]glycophorin A. 13C NMR spectra of these species show that the resonances of the methyl groups of the modified glycophorins occur at 26.1 ppm downfield from Me4Si. A spin-lattice relaxation time of 0.4 s was observed for the 13C-enriched methyl resonances of the sulfonium ion derivatives of Met-8 and -81, which corresponds to an effective correlation time of less than 2 X 10 - 10 s. Demethylation of the 2 glycophorin A sulfonium ion species with 2-mercaptoethanol produces native glycophorin A which now has the epsilon-carbon of the methionine residue(s) 45% isotopically enriched. The epsilon-carbon of Met-8 was found to occur at 15.7 ppm downfield from Me4Si whereas the epsilon-carbon of Met-81 exhibited an unusual chemical shift of 2.0 ppm downfield from Me4Si. The spin-lattice relaxation time of both resonances was found to be approximately 0.3 s.