title: The role of backbone hydrogen bonds in the transition state for protein folding of a PDZ domain.
abstract: Backbone hydrogen bonds are important for the structure and stability of proteins. However, since conventional site-directed mutagenesis cannot be applied to perturb the backbone, the contribution of these hydrogen bonds in protein folding and stability has been assessed only for a very limited set of small proteins. We have here investigated effects of five amide-to-ester mutations in the backbone of a PDZ domain, a 90-residue globular protein domain, to probe the influence of hydrogen bonds in a β-sheet for folding and stability. The amide-to-ester mutation removes NH-mediated hydrogen bonds and destabilizes hydrogen bonds formed by the carbonyl oxygen. The overall stability of the PDZ domain generally decreased for all amide-to-ester mutants due to an increase in the unfolding rate constant. For this particular region of the PDZ domain, it is therefore clear that native hydrogen bonds are formed after crossing of the rate-limiting barrier for folding. Moreover, three of the five amide-to-ester mutants displayed an increase in the folding rate constant suggesting that the hydrogen bonds are involved in non-native interactions in the transition state for folding.
authors.affiliation: Department of Medical Biochemistry and Microbiology, Uppsala University, Biomedical Center, Uppsala, Sweden; Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Søren W
authors.initials: SW
authors.lastname: Pedersen
authors.affiliation: Department of Medical Biochemistry and Microbiology, Uppsala University, Biomedical Center, Uppsala, Sweden.
authors.forename: Greta
authors.initials: G
authors.lastname: Hultqvist
authors.affiliation: Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Kristian
authors.initials: K
authors.lastname: Strømgaard
authors.affiliation: Department of Medical Biochemistry and Microbiology, Uppsala University, Biomedical Center, Uppsala, Sweden.
authors.forename: Per
authors.initials: P
authors.lastname: Jemth
mesh_terms: D002942:Circular Dichroism; D006860:Hydrogen Bonding; D007700:Kinetics; D008958:Models, Molecular; D054731:PDZ Domains; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry*; D013816:Thermodynamics
chemical_list: D011506:Proteins
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