title: Crystal structures of two mutants that have implications for the folding of bovine pancreatic ribonuclease A.
abstract: The Tyr92-Pro93 peptide group of bovine pancreatic ribonuclease A (RNase A) exists in the cis conformation in the native state. From unfolding/refolding kinetic studies of the disulfide-intact wild-type protein and of a variant in which Pro93 had been replaced by Ala, it had been suggested that the Tyr92-Ala93 peptide group also exists in the cis conformation in the native state. Here, we report the crystal structure of the P93A variant. Although there is disorder in the region of residues 92 and 93, the best structural model contains a cis peptide at this position, lending support to the results of the kinetics experiments. We also report the crystal structure of the C[40, 95]A variant, which is an analog of the major rate-determining three-disulfide intermediate in the oxidative folding of RNase A, missing the 40-95 disulfide bond. As had been detected by NMR spectroscopy, the crystal structure of this analog shows disorder in the region surrounding the missing disulfide. However, the global chain fold of the remainder of the protein, including the disulfide bond between Cys65 and Cys72, appears to be unaffected by the mutation.
authors.affiliation: Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853-1301, USA.
authors.forename: M A
authors.initials: MA
authors.lastname: Pearson
authors.forename: P A
authors.initials: PA
authors.lastname: Karplus
authors.forename: R W
authors.initials: RW
authors.lastname: Dodge
authors.forename: J H
authors.initials: JH
authors.lastname: Laity
authors.forename: H A
authors.initials: HA
authors.lastname: Scheraga
mesh_terms: D000818:Animals; D002417:Cattle; D018360:Crystallography, X-Ray; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D009154:Mutation; D010179:Pancreas / Q000201:enzymology*; D011487:Protein Conformation; D017510:Protein Folding; D012259:Ribonuclease, Pancreatic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D012259:Ribonuclease, Pancreatic