Solvent Proton Magnetic Relaxation Dispersion in Solutions of Concanavalin A

Abstract

Concanavalin A, a protein isolated from jack beans, exhibits several important biological properties, all of which are related to its ability to bind and precipitate specific polysaccharides. Concanavalin A is a dimer at pH 5.6, and has one transition-metal and one calcium-ion binding site per monomer unit of molecular weight 27,000. Both metal-ion sites must be occupied for the protein to be active. It is of interest to determine the role of the transition metal ion in Concanavalin A and its relationship to the sugar binding activity of the protein.We report the magnetic field and temperature dependences of the spin-lattice magnetic relaxation rates of solvent protons in aqueous solutions of zinc and manganese derivatives of Concanavalin A, and the influence of monosaccharide binding on these rates. The results of a leastsquares fit of the data to the theory, with five adjustable parameters, indicate that there is one rapidly exchanging water molecule ligand on the Mn2+ ion, with a residence lifetime of 2.5 gsec at 250, and with its protons 0.27 nm (2.7 A) from the Mn2+ ion. We find that at low magnetic fields (proton Larmor frequencies below about 10 MHz), the correlation time for the dipolar interaction between the Mn2+ electronic spin moment and the protons on the water ligand is the spin-lattice relaxation time rs of the Mn2+ moment, but that at higher magnetic fields the correlation time for the dipolar interaction is determined by the Brownian rotational tumbling of the protein, because of the substantial variation of TS with magnetic field. Monosaccharide binding to manganese Concanavalin A has little effect on the relaxation rates of solvent protons, a result that indicates that the sugars do not bind directly to the transition metal in the protein.The jack-bean protein Concanavalin A (Con A), a member of the lectin class of plant proteins, possesses hemagglutinating activity (1) and precipitates various polysaccharides, such as glycogen and yeast mannans (2), in a manner analogous to antibody-antigen reactions. The protein also specifically binds monosaccharides such as a-methyl-D-glucopyranoside (3). Among the more intriguing biological properties of Con A is its ability to agglutinate leukemic cells and cells transformed by polyoma virus or simian virus 40 (4). Con A (treated with trypsin) also returns transformed fibroblasts in tissue cultures to normal growth behavior (5) and inhibits various tumors in vivo (6). In addition, the protein agglutinates epithelial cells isolated from the intestine of human fetuses, but not those of Abbreviations: Con A, Concanavalin A; ESR, electron spin resonance; NMR, nuclear magnetic resonance. * This is paper VI in a series on solvent proton relaxation dis- adults (7). The addition of competitively binding monosaccharides reverses these biological effects of Con A (4-7).Con A is a dimer between pH 3.5 and 5.6, with a molecular weight of 54,000; higher polymeric species form above pH 5.6 (8, 9). Each monomer has a binding site for transition metal ions wh...

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