Fructose-1,6-bisphosphate-activated pyruvate kinase from Escherichia coli. Nature of bonds involved in the allosteric mechanism.
The allosteric properties of the fructose-1,6-bis-phosphate-activated pyruvate kinase from Escherichia coli were examined in the presence of a number of fructose bisphosphate analogues, as well as of increased ionic strength (NaCl) and of the hydrogen-bond-breaking agent, formamide. Fructose 2,6-bisphosphate, ribulose 1,5-bisphosphate and 5-phosphorylribose 1-pyrophosphate gave allosteric activation (additive to that of fructose 1,6-bisphosphate). Formamide always decreased Vmax, but left unchanged the Km for phosphoenolpyruvate, while it decreased the concentration of fructose bisphosphate required to give half-maximal activity (K0.5). NaCl increased the K0.5 for both phosphoenolpyruvate and fructose bisphosphate, leaving Vmax unchanged. These results are consistent with ionic binding of fructose bisphosphate through phosphates and with a critical role of hydrogen bonds in stabilizing both the inactive and the active enzyme conformers.
Speranza
M L
ML
Dipartimento di Biochimica, Università di Pavia, Italy.
Valentini
G
G
Malcovati
M
M
D000494:Allosteric Regulation; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000201:enzymology*; D005559:Formamides / Q000494:pharmacology; D005635:Fructosediphosphates / Q000494:pharmacology*; D006598:Hexosediphosphates / Q000494:pharmacology*; D006860:Hydrogen Bonding; D007700:Kinetics; D015394:Molecular Structure; D009994:Osmolar Concentration; D010728:Phosphoenolpyruvate / Q000378:metabolism; D011770:Pyruvate Kinase / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology
D005559:Formamides; D005635:Fructosediphosphates; D006598:Hexosediphosphates; D012965:Sodium Chloride; C031066:formamide; D010728:Phosphoenolpyruvate; D011770:Pyruvate Kinase; C029063:fructose-1,6-diphosphate