Related Experiment Video
Updated: Sep 3, 2025
Measuring In Vitro ATPase Activity for Enzymatic Characterization
Published on: August 23, 2016
Replacing Commercial 6-Phosphofructokinase in an Online Pyrophosphate Detection Assay
Daniel L Machell1,2,3, Mathias H Hansen1,2,3, Max J Cryle1,2,3
1The Monash Biomedicine Discovery Institute, Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, 3800, Australia.
Abstract:
Detection of pyrophosphate is important in quantifying enzyme activity, particularly adenylation domain activity during non-ribosomal peptide synthesis. The previous development of an enzyme coupled PPi /NADH assay allowed the measurement of such activity in an online fashion using commercially available components. Now, with a key enzyme - 6-phosphofructokinase - no longer available, we have screened and identified viable replacement enzymes that can be expressed in high yield and that are far superior in activity to the now discontinued commercial product. This will support the ability of groups to continue to use this established online assay for pyrophosphate detection.
