Orientational order and dynamics of hydration water in a single crystal ofbovine pancreatic trypsin inhibitor

The orientational order and dynamics of the water molecules in form II crys tals of bovine pancreatic trypsin inhibitor (BPTI) are studied by H-2 NMR i n the temperature range 6-50 degrees C. From the orientation dependence of the single crystal quadrupole splitting and linewidth, the principal compon ents of the motionally averaged quadrupole interaction tensor and the irred ucible linewidth components for the orthorhombic crystal are determined. Wi th the aid of water orientations derived from neutron and x-ray diffraction , it is shown that the NMR data can be accounted for by a small number of h ighly ordered crystal waters, some of which have residence times in the mic rosecond range. Most of these specific hydration sites must be located at i ntermolecular contacts. The surface hydration layer that is also present in dilute solution is likely to be only weakly ordered and would then not con tribute significantly to the splining and linewidth from the protein crysta l. To probe water dynamics on shorter time scales, the 2H longitudinal rela xation dispersion is measured for a polycrystalline BPTI sample. The observ ed dispersion is dominated by rapidly exchanging deuterons in protein side chains, undergoing restricted rotational motions on a time scale of 10 ns.