The binding of glucose and nucleotides to hexokinase from Saccharomyces cerevisiae (original) (raw)

1988, Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology

The binding of glucose, ADP and AdoPPINH]P, to the native PII dimer and PII monomer and the proteolytically-modified SII monomer of hexokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1) from Saccharomyces cerevisiae was monitored at pH 6.7 by the concomitant quenching of protein fluorescence. The data were analysed in terms of Qm~,, the maximal quenching of fluorescence at saturating concentrations of ligand, and [L]0.s, the concentration of ligand at half-maximal quenching. No changes in fluorescence were observed with free enzyme and nucleotide alone. In the presence of saturating levels of glucose, Qmax induced by nucleotide was between 2 and 7%, and [L]0. 5 was between 0.12 and 0.56 mM, depending on the nucleotide and enzyme species. Omax induced by glucose alone was between 22 and 25%, while [L] 0.5 was approx. 0.4 mM for either of the monomeric hexokinase forms and 3.4 for PII dimer. In the presence of 6 mM ADP or 2 mM AdoPP[NH]P, Qm~ for glucose was increased by up to 4% and [L]0. s was diminished 3-fold for hexokinase PII monomer, 6-fold for SII monomer, and 15-fold for PII dimer. The results are interpreted in terms of nucleotide-induced conformational change of bexokinase in the presence of glucose and synergistic binding interactions between glucose and nucleotide.

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