A sequential binding mechanism in a PDZ domain

Research output: Contribution to journalJournal articleResearchpeer-review

Celestine N Chi, Anders Bach, Åke Engström, Huiqun Wang, Kristian Strømgaard, Stefano Gianni, Per Jemth

Conformational selection and induced fit are two well-known mechanisms of allosteric protein-ligand interaction. Some proteins, like ubiquitin, have recently been found to exist in multiple conformations at equilibrium, suggesting that the conformational selection may be a general mechanism of interaction, in particular for single-domain proteins. Here, we found that the PDZ2 domain of SAP97 binds its ligand via a sequential (induced fit) mechanism. We performed binding experiments using SAP97 PDZ2 and peptide ligands and observed biphasic kinetics with the stopped-flow technique, indicating that ligand binding involves at least a two-step process. By using an ultrarapid continuous-flow mixer, we then detected a hyperbolic dependence of binding rate constants on peptide concentration, corroborating the two-step binding mechanism. Furthermore, we found a similar dependence of the rate constants on both PDZ and peptide concentration, demonstrating that the PDZ2-peptide interaction involves a precomplex, which then undergoes a conformational change, and thereby follows an induced fit mechanism.
Original languageEnglish
JournalBiochemistry
Volume48
Issue number30
Pages (from-to)7089-7097
ISSN0006-2960
DOIs
Publication statusPublished - 2009

Bibliographical note

Keywords: Adaptor Proteins, Signal Transducing; Amino Acid Sequence; Binding Sites; DNA-Binding Proteins; Humans; Ligands; Membrane Proteins; Models, Molecular; Molecular Sequence Data; Oncogene Proteins, Viral; PDZ Domains; Peptides; Protein Binding; Protein Conformation; Protein Denaturation; Protein Folding; Thermodynamics

ID: 17365971