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Updated: Aug 10, 2026
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Direct Detection of the Acetate-forming Activity of the Enzyme Acetate Kinase
Published on: December 19, 2011
Allosteric activation increases the maximum velocity of E. coli phosphofructokinase
I Auzat1, G Le Bras, J R Garel
1Laboratoire d'Enzymologie et de Biochimie Structurales, Gif-sur-Yvette, France.
Journal of Molecular Biology
|April 4, 1997
Summary
GDP activation of phosphofructokinase involves changes in maximum velocity, not just substrate affinity. Mutations reveal GDP
Area of Science:
- Biochemistry
- Enzymology
- Molecular Biology
Background:
- Phosphofructokinase (PFK) is a key glycolytic enzyme regulated by allosteric activators.
- Guanosine diphosphate (GDP) is known to activate Escherichia coli PFK, influencing its kinetics.
Purpose of the Study:
- To investigate the role of GDP as an allosteric activator on wild-type and mutant phosphofructokinase from Escherichia coli.
- To elucidate the specific effects of GDP on enzyme kinetics, including cooperativity and maximum velocity.
Main Methods:
- Site-directed mutagenesis was used to introduce specific amino acid substitutions in E. coli phosphofructokinase.
- Enzyme kinetics were measured for wild-type and mutant enzymes in the presence and absence of GDP.
- Saturation kinetics with fructose-6-phosphate were analyzed to determine cooperativity and affinity changes.
Main Results:
- Mutants Glu148 --> Leu, Leu178 --> Val, and Leu178 --> Trp retained cooperativity, which was reduced by GDP. GDP increased the maximum velocity of these mutants by 35-65%.
Conclusions:
- GDP's activation of wild-type phosphofructokinase may involve an increase in maximum velocity, not solely an enhancement of fructose-6-phosphate affinity.
- Specific mutations impact GDP's allosteric regulation, highlighting the interplay between enzyme structure and activator function.
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