title: Mutations that destabilize the a' domain of human protein-disulfide isomerase indirectly affect peptide binding.
abstract: Protein-disulfide isomerase (PDI) is a catalyst of folding of disulfide-bonded proteins and also a multifunctional polypeptide that acts as the beta-subunit in the prolyl 4-hydroxylase alpha(2)beta(2)-tetramer (P4H) and the microsomal triglyceride transfer protein alphabeta-dimer. The principal peptide-binding site of PDI is located in the b' domain, but all domains contribute to the binding of misfolded proteins. Mutations in the C-terminal part of the a' domain have significant effects on the assembly of the P4H tetramer and other functions of PDI. In this study we have addressed the question of whether these mutations in the C-terminal part of the a' domain, which affect P4H assembly, also affect peptide binding to PDI. We observed a strong correlation between P4H assembly competence and peptide binding; mutants of PDI that failed to form a functional P4H tetramer were also inactive in peptide binding. However, there was also a correlation between inactivity in these assays and indicators of conformational disruption, such as protease sensitivity. Peptide binding activity could be restored in inactive, protease-sensitive mutants by selective proteolytic removal of the mutated a' domain. Hence we propose that structural changes in the a' domain indirectly affect peptide binding to the b' domain.
authors.affiliation: Department of Biosciences, University of Kent, Canterbury CT2 7NJ, United Kingdom. P.Klappa@ukc.ac.uk
authors.forename: P
authors.initials: P
authors.lastname: Klappa
authors.forename: P
authors.initials: P
authors.lastname: Koivunen
authors.forename: A
authors.initials: A
authors.lastname: Pirneskoski
authors.forename: P
authors.initials: P
authors.lastname: Karvonen
authors.forename: L W
authors.initials: LW
authors.lastname: Ruddock
authors.forename: K I
authors.initials: KI
authors.lastname: Kivirikko
authors.forename: R B
authors.initials: RB
authors.lastname: Freedman
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases / Q000737:chemistry* / Q000235:genetics
chemical_list: D010455:Peptides; D019704:Protein Disulfide-Isomerases