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Assembly of PGAM5 into Multimeric Complexes Provides a Mechanism for Allosteric Regulation of Phosphatase Activity

Peter Tipton1, Tong Su1, Mark Hannink1

  • 1University of Missouri, Columbia, MO, United States.

Methods in Enzymology
|August 29, 2018
PubMed

Insights

Phosphoglycerate mutase family member 5 (PGAM5) is a mitochondrial phosphatase regulating cell physiology. This study details methods to purify PGAM5 complexes and characterize their varying phosphatase activities for drug screening.

Area of Science:

  • Mitochondrial biology
  • Enzymology
  • Cellular signaling

Background:

  • Phosphoglycerate mutase family member 5 (PGAM5) is a mitochondrial serine/threonine phosphatase.
  • PGAM5 influences mitochondrial dynamics, including fission/fusion and mitophagy, via distinct mechanisms.
  • Understanding PGAM5 regulation is crucial for linking cellular physiology to mitochondrial function.

Purpose of the Study:

  • To describe methods for purifying multimeric and dimeric PGAM5 complexes.
  • To characterize the kinetic properties of different PGAM5 complexes.
  • To enable biophysical studies of PGAM5 quaternary structures and facilitate modulator screens.

Main Methods:

  • Purification of multimeric and dimeric PGAM5 complexes.
  • Kinetic characterization of purified PGAM5 complexes.
  • Biophysical analysis of PGAM5 quaternary structures.

Main Results:

  • PGAM5 complexes purified in distinct multimeric and dimeric forms.
  • Phosphatase activity of PGAM5 complexes varies significantly across three orders of magnitude.
  • Purified complexes enable detailed biophysical and kinetic studies.

Conclusions:

  • Methods allow for the purification and characterization of PGAM5 complexes with diverse activities.
  • This facilitates biophysical investigations into PGAM5 structure-function relationships.
  • Enables screening for small molecules that modulate PGAM5 phosphatase activity for therapeutic potential.
Keywords:
Allosteric regulationHyperbolic noncompetitive inhibitionMultimer complex assemblyPGAM5Quaternary structureRapid equilibrium-ordered mechanism

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