Temperature dependence of binding and catalysis for the Cdc25B phosphatase

Jungsan Sohn, Johannes Rudolph

Research output: Contribution to journalArticlepeer-review

6 Scopus citations


Using a combination of steady-state and single-turnover kinetics, we probe the temperature dependence of substrate association and chemistry for the reaction of Cdc25B phosphatase with its Cdk2-pTpY/CycA protein substrate. The transition state for substrate association is dominated by an enthalpic barrier (ΔH of 13 kcal/mol) and has a favorable entropic contribution of 4 kcal/mol at 298 K. Phosphate transfer from Cdk2-pTpY/CycA to enzyme (ΔH of 12 kcal/mol) is enthalpically more favorable than for the small molecule substrate p-nitrophenyl phosphate (ΔH of 18 kcal/mol), yet entropically less favorable (TΔS of 2 vs. - 6 kcal/mol at 298 K, respectively). By measuring the temperature dependence of binding and catalysis for several hotspot mutants involved in binding of protein substrate, we determine the enthalpy-entropy compensations for changes in rates of association and phosphate transfer compared to the wild type system. We conclude that the transition state for enzyme-substrate association involves tight and specific contacts at the remote docking site and that phospho-transfer from Cdk2-pTpY/CycA to the pre-organized active site of the enzyme is accompanied by unfavorable entropic rearrangements that promote rapid product dissociation.

Original languageEnglish (US)
Pages (from-to)549-555
Number of pages7
JournalBiophysical Chemistry
Issue number2-3
StatePublished - Feb 2007
Externally publishedYes


  • Protein tyrosine phosphatase
  • Protein-protein interactions
  • Single-turnover kinetics
  • Thermodynamics

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Organic Chemistry


Dive into the research topics of 'Temperature dependence of binding and catalysis for the Cdc25B phosphatase'. Together they form a unique fingerprint.

Cite this