Pyruvate kinase of Streptococcus lactis

Area of Science:

• Biochemistry
• Enzymology
• Microbial Metabolism

Background:

• Pyruvate kinase (ATP:pyruvate-phosphotransferase, EC 2.7.1.40) is a crucial enzyme in glycolysis.
• Understanding its kinetic properties is vital for elucidating metabolic regulation in microorganisms.

Purpose of the Study:

• To investigate the kinetic properties of pyruvate kinase from Streptococcus lactis.
• To determine the effects of substrates and allosteric effectors on enzyme activity.

Main Methods:

• Enzyme kinetics assays were performed.
• Kinetic parameters (Km, Vmax) were determined under varying substrate and effector concentrations.
• Effect of pH and inhibitors were assessed.

Main Results:

• Pyruvate kinase displayed positive homotropic kinetics with phosphoenolpyruvate and ADP, showing sigmoid substrate-velocity curves.
• Fructose-1,6-diphosphate acted as a heterotropic activator, converting kinetics to Michaelis-Menten type and increasing Vmax while decreasing Km.
• Optimal activity was observed at pH 6.9-7.5, with inhibition by inorganic phosphate, sulfate, and under certain conditions, ATP.
• Intracellular metabolite concentrations suggest near-maximal enzyme activity in growing S. lactis cells.

Conclusions:

• Streptococcus lactis pyruvate kinase is allosterically regulated by fructose-1,6-diphosphate.
• The enzyme's kinetic properties are consistent with its role in regulating glycolysis in vivo.
• This regulation is crucial for energy metabolism in lactic streptococci.