Deleterious effects of beta-branched residues in the S-1 specificity pocket of Streptomyces griseus proteinase B (SGPB): Crystal structures of the turkey ovomucoid third domain variants Ile18I, Val18I, Thr18I, and Ser18I in complex with SGPB

Turkey ovomucoid third domain (OMTKY3) is a canonical inhibitor of serine p roteinases. Upon complex formation, the inhibitors fully exposed P-1 residu e becomes fully buried in the preformed cavity of the enzyme. All 20 P-1 va riants of OMTKY3 have been obtained by recombinant DNA technology and their equilibrium association constants have been measured with six serine prote inases. To rationalize the trends observed in this data set, high resolutio n crystal structures have been determined for OMTKY3 P-1 variants in comple x with the bacterial serine proteinase, Streptomyces griseus proteinase B ( SGPB). Four high resolution complex structures are being reported in this p aper; the three beta-branched variants, Ile18I, Val18I, and Thr18I, determi ned to 2.1, 1.6, and 1.7 Angstrom resolution, respectively, and the structu re of the Ser18I variant complex, determined to 1.9 Angstrom resolution. Mo dels of the Cys18I, Hse18I, and Ape18I variant complexes are also discussed . The beta-branched side chains are not complementary to the shape of the S -1 binding pocket in SGPB, in contrast to that of the wild-type gamma-branc hed P-1 residue for OMTKY3, Leu18I. chi(1) angles of approximately 40 degre es are imposed on the side chains of Ile18I, Val18I, and Thr18I within the S-1 pocket. Dihedral angles of +60 degrees, -60 degrees, or 180 degrees are more commonly observed but 40 degrees is not unfavorable for the beta-bran ched side chains. Thr18I O-gamma 1 also forms a hydrogen bond with Ser195 O -gamma in this orientation. The Ser18I side chain adopts two alternate conf ormations within the S-1 pocket of SGPB, suggesting that the side chain is not stable in either conformation.