Cryocrystallography of a Kunitz-type serine protease inhibitor: the 90 K structure of winged bean chymotrypsin inhibitor (WCI) at 2.13 angstrom resolution

The crystal structure of a Kunitz-type double-headed alpha-chymotrypsin inh ibitor from winged bean seeds has been refined at 2.13 Angstrom resolution using data collected from cryo-cooled (90 K) crystals which belong to the h exagonal space group P6(1)22 with unit-cell parameters a = b = 60.84, c = 2 07.91 Angstrom. The volume of the unit cell is reduced by 5.3% on cooling. The refinement converged to an R value of 20.0% (R-free = 25.8%) for 11100 unique reflections and the model shows good stereochemistry, with r.m.s. de viations from ideal values for bond lengths and bond angles of 0.011 Angstr om and 1.4 degrees, respectively. The structural architecture of the protei n consists of 12 antiparallel beta-strands joined in the form of a characte ristic beta-trefoil fold, with the two reactive-site regions, Asn(38)-Leu43 and Gln63-Phe68, situated on two external loops. Although the overall prot ein fold is the same as that of the room-temperature model, some conformati onal changes are observed in the loop regions and in the side chains of a f ew surface residues. A total of 176 ordered water molecules and five sulfat e ions are included in the model.