Functional energetic landscape in the allosteric regulation of muscle pyruvate kinase. 1. Calorimetric study

Petr Herman, J. Ching Lee

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

Rabbit muscle pyruvate kinase (RMPK) is an important allosteric enzyme of the glycolytic pathway catalyzing a transfer of the phosphate from phosphoenolpyruvate (PEP) to ADP. The energetic landscape of the allosteric regulatory mechanism of RMPK was characterized by isothermal titration calorimetry (ITC) in the temperature range from 4 to 45°C. ITC data for RMPK binding to substrates PEP and ADP, for the allosteric inhibitor Phe, and for combination of ADP and Phe were globally analyzed. The thermodynamic parameters characterizing the linked-multiple-equilibrium system were extracted. Four novel insights were uncovered. (1) The binding preference of ADP for either the T or R state is temperature-dependent, namely, more favorable to the T and R states at high and low temperatures, respectively. This crossover of affinity toward R and T states implies that ADP plays a complex role in modulating the allosteric behavior of RMPK. Depending on the temperature, binding of ADP can regulate RMPK activity by favoring the enzyme to either the Ror T state. (2) The binding of Phe is negatively coupled to that of ADP; i.e., Phe and ADP prefer not to bind to the same subunit of RMPK. (3) The release or absorption of protons linked to the various equilibria is specific to the particular reaction. As a consequence, pH will exert a complex effect on these linked equilibria, resulting in the proton being an allosteric regulatory ligand of RMPK. (4) The R ↔ T equilibrium is accompanied by a significant ΔCp, rendering RMPK most sensitive to temperature under physiological conditions. During muscle activity, both pH and temperature fluctuations are known to happen; thus, results of this study are physiologically relevant.

Original languageEnglish (US)
Pages (from-to)9448-9455
Number of pages8
JournalBiochemistry
Volume48
Issue number40
DOIs
StatePublished - 2009

ASJC Scopus subject areas

  • Biochemistry

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