Magnetic resonance studies of concanavalin A: location of the binding site of alpha-methyl-D-mannopyranoside.

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Abstract
The longitudinal nuclear magnetic relaxation times of the methyl protons of alpha-methyl-D-mannopyranoside have been measured at 90 MHz and 270 MHz in solutions of concanavalin A complexed with: (i) Mn2+ and Ca2+; and (ii) Zn2+ and Ca2+. Zn2+ and Mn2+ are known to bind in site S1 and Ca2+ in site S2 of concanavalin A. Both sites must be occupied before monosaccharides will bind to the protein. In order to extract T1p, the paramagnetic contribution to the bound methyl relaxation time, from these observations the relaxation time of uncomplexed sugar was determined. Free Mn2+ contributed insignificantly to the latter value; however, outer sphere relaxation was found to be large at bound Mn2+ concentrations greater than 1 mM. Comparison of the results obtained at the two frequencies allowed the determination of the correlation time for the interaction between the methyl protons and the bound Mn2+ and the distance between them (21.5 +/- 1.2 A). In contrast to previous results from nuclear magnetic resonance studies, this distance is consistent with binding at the cavity proposed for the saccharide binding site by Becker et al. [J. Biol. Chem. 250, 1513 (1975)], although it does not preclude possible binding sites on the surface of the molecule.