Hydration and activity studies of bovine skeletal muscle proteins and wheat gliadins by nuclear magnetic resonance

Abstract

\sp1H and \sp{17}O NMR studies of water and ion binding to bovine skeletal muscle proteins in solutions and suspensions were carried out as a function of concentration, pH/pD, and salt. Numerical analysis of the data was performed by nonlinear regression on an Apple-MacIntosh II microcomputer, using a thermodynamic linkage approach. The second virial coefficient, B\sb{\rm o}, of myosin $A$ and $B$ was obtained from the protein concentration dependence of the relaxation rates at high ionic strength (with 0.5 M NaCl). Myosin self-association in the form of tail-to-tail dimers was inferred from the NMR data in the absence of salt. At higher salt concentrations than 0.6 M NaCl, heptamer formation was inferred for myofibrillar proteins, whereas in the presence of 0.5 M NaCl, myosin was found to be fully dispersed as monomers. The "binding" of water to myosin $A$ appears to involve primarily carboxyl groups at high pH and imidazolium or ammonium groups at low pH, with hydration being minimal at the isoelectric point (5.5). The molecular dynamics of hydration water and ions bound to myosin occurred on a time scale of tens of picoseconds. The analysis of \sp{23}Na NMR studies of Na\sp+ binding to myofibrillar proteins reveals the presence of two types of binding sites on these proteins.Binding of Mn\sp{2+} ions to wheat gliadins was studied by observing the paramagnetic relaxation enhancement of the "bound" water protons at 10 and 20 MHz; from such studies the number of charged carboxyl groups was found to be n = 7 $\pm$ 1 at pH 3.4 and 25\sp\circC, with a binding constant for Mn\sp{2+}, K\sb{\rm f} = 12.9 M\sp{-1}. \sp\tau\sb{\rm c} of water "bound" to Mn\sp{2+} attached to wheat gliadins in solutions was determined to be 18 ns.U of I OnlyETDs are only available to UIUC Users without author permissio