Physical studies of 13C-methylated concanavalin A. pH- and Co2+-induced nuclear magnetic resonance shifts.

A. D. Sherry, J. Teherani

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19 Scopus citations

Abstract

Concanavalin A has been reductively methylated using [13C]formaldehyde and NaCNBH3 and examined by 13C NMR. The chemical modification does not alter the metal ion or saccharide binding properties of the protein nor the associating of dimers to form tetramers above pH 6. Eleven of the 12 N epsilon,N-dimethyllysines appear as a broad unresolved group of resonances at pH 5.6, while one N epsilon,N-dimethyllysine (tentatively assigned to Lys 101) gives rise to two resolved methyl resonances as a result of forming a salt bridge with Asp 203 (Reeke, G. N., Jr., Becker, J. W., and Edelman, G. M. (1975) J. Biol. Chem. 250, 1525-1547). The NH2-terminal N alpha,N-dimethylalanine appears as a unique resonance and titrates with a pKa of 7.9. The chemical shift degeneracy of the N epsilon,N-dimethyllysine is lifted when Co2+ is bound in the S1 site of the protein. These resonances sharpen and shift downfield differentially with increasing pH until eight of 12 N epsilon,N-dimethyllysine resonances are resolved at pH 10. A comparison of the expected Co2+-induced shift at each lysine based upon their crystal coordinates and previous Co2+-histidine shift data (Carver, J. P., Barber, B. H., and Fuhr, B. J. (1977) J. Biol. Chem. 252, 3141-3146) with those observed in the N epsilon,N-dimethyllysine resonances has allowed a tentative assignment of several resonances to a specific lysine in the sequence.

Original languageEnglish (US)
Pages (from-to)8663-8669
Number of pages7
JournalJournal of Biological Chemistry
Volume258
Issue number14
StatePublished - Jul 25 1983

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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