Intracellular hexose-6-phosphate:phosphohydrolase from Streptococcus lactis: purification, properties, and function.

Area of Science:

• Biochemistry
• Enzymology
• Microbial Metabolism

Background:

• Streptococcus lactis K1 possesses an intracellular hexose 6-phosphate:phosphohydrolase (EC 3.1.3.2).
• This enzyme plays a role in the phosphoenolpyruvate-dependent futile cycle.

Purpose of the Study:

• To purify and characterize the intracellular hexose 6-phosphate:phosphohydrolase from Streptococcus lactis K1.
• To elucidate the enzyme's substrate specificity and cofactor requirements.

Main Methods:

• Enzyme purification using polyacrylamide disc gel electrophoresis and gel permeation chromatography.
• Substrate specificity determined by measuring hydrolysis rates of various phosphorylated compounds.
• Enzyme activity assays with different divalent metal ions and inhibitors.

Main Results:

• Purified enzyme showed one major activity staining protein band with an apparent Mr of 60,000.
• Enzyme exhibited preference for galactose 6-phosphate and 2-deoxy-D-glucose 6-phosphate.
• Activity required Mn2+, Mg2+, Fe2+, or Co2+ for activation; Pb2+, Cu2+, Zn2+, Cd2+ were inhibitors.
• Fluoride inhibition was observed with Mg2+ but not Mn2+, Fe2+, or Co2+.

Conclusions:

• The purified hexose 6-phosphate:phosphohydrolase is crucial for catalyzing 2-deoxy-D-glucose 6-phosphate hydrolysis in S. lactis.
• Metal ion cofactors significantly influence enzyme activity and inhibition patterns.
• Understanding this enzyme provides insight into microbial metabolic pathways.