Proton and Deuteron Nuclear Magnetic Relaxation Dispersion Studies of Ca²+-Mn²+-Lentil Lectin and Ca²⁺-Mn²⁺-Pea Lectin: Evidence for a Site of Solvent Exchange in Common with Concanavalin A

Measurements of the magnetic field dependence of the longitudinal magnetic relaxation rates (NMRD profiles) of solvent protons and deuterons led to the discovery of two classes of solvent binding sites in Ca²⁺-Mn²⁺-concanavalin A (CMPL) [Koenig, S. H., Brown, R. D., III, & Brewer, C. F. (1985) Biochemistry (second of three papers in this issue)]. In this paper, we compare proton and deuteron NMRD profiles of Ca²⁺-Mn²⁺-lentil lectin (CMLcH) and Ca²⁺-Mn²⁺-pea lectin (CMPSA) with those of CMPL. All three metalloproteins are d-mannose/d-glucose-specific lectins that have a high degree of structural similarity and require the metal ions for their biological activities. We have developed a method for the preparation of fully active metal ion derivatives of lentil lectin (LcH) and pea lectin (PSA), including the diamagnetic derivatives Ca2+-Zn2+-LcH and Ca2+-Zn2+-PSA [Bhattacharyya, L., Brewer, C. F., Brown, R. D., III, & Koenig, S. H.(1984) Biochem. Biophys. Res. Commun. 124, 857-862]. The behavior of these two lectins with regard to their NMRD profiles is essentially identical, for both the paramagnetic and diamagnetic forms. Together with CMPL, all three lectins have a common paramagnetic contribution with a negative temperature dependence of the rates, while CMPL contributes an additional component with a positive temperature dependence. The common contribution derives from the class of fast exchanging water molecules observed in the proton NMRD profile of CMPL (Koenig et al., 1985); their protons are calculated to be relatively remote from the Mn²⁺ ions (4.4 Å for CMPL and 5.5 Å for LcH and PSA). These waters are tentatively assigned to the coordination sphere of the Ca²⁺ ions in all three proteins, and their relaxation contributions are insensitive to the presence of bound saccharide.