A sequential binding mechanism in a PDZ domain
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A sequential binding mechanism in a PDZ domain. / Chi, Celestine N; Bach, Anders; Engström, Åke; Wang, Huiqun; Strømgaard, Kristian; Gianni, Stefano; Jemth, Per.
I: Biochemistry, Bind 48, Nr. 30, 2009, s. 7089-7097.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - A sequential binding mechanism in a PDZ domain
AU - Chi, Celestine N
AU - Bach, Anders
AU - Engström, Åke
AU - Wang, Huiqun
AU - Strømgaard, Kristian
AU - Gianni, Stefano
AU - Jemth, Per
N1 - 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
PY - 2009
Y1 - 2009
N2 - 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.
AB - 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.
KW - Former Faculty of Pharmaceutical Sciences
U2 - 10.1021/bi900559k
DO - 10.1021/bi900559k
M3 - Journal article
C2 - 19496620
VL - 48
SP - 7089
EP - 7097
JO - Biochemistry
JF - Biochemistry
SN - 0006-2960
IS - 30
ER -
ID: 17365971