PLANT PHYSIOLOGY , Vol 103, Issue 1 285-288, Copyright © 1993 by American Society of Plant Biologists
Activation of Cytosolic Pyruvate Kinase by Polyethylene Glycol
F. E. Podesta and W. C. Plaxton
Departments of Biology and Biochemistry, Queen's University, Kingston, Ontario, Canada K7L 3N6
Homogeneous cytosolic pyruvate kinase from endosperm of germinating castor
oil (Ricinus communis L. cv Hale) seeds was potently activated by
polyethylene glycol. The addition of 5% (w/v) polyethylene glycol to the
pyruvate kinase reaction mixture caused a 2.6-fold increase in maximal
velocity and 12.5- and 2-fold reductions in Km values for
phosphoenolpyruvate and ADP, respectively. Glycerol, ethylene glycol, and
bovine serum albumin also enhanced pyruvate kinase activity, albeit to a
lesser extent than polyethylene glycol. The addition of 5% (w/v)
polyethylene glycol to the elution buffer during high-performance gel
filtration chromatography of purified cytosolic pyruvate kinase helped to
stabilize the active heterotetrameric native structure of the enzyme. A
higher degree of inhibition by MgATP, but lower sensitivity to the
inhibitors 3-phosphoglycerate and fructose- 1,6-bisphosphate, was also
observed in the presence of 5% (w/v) polyethylene glycol. It is concluded
that (a) plant cytosolic pyruvate kinase activity and regulation, like that
of other regulatory pyruvate kinases, is modified by extreme dilution in
the assay medium, probably as a result of deaggregation of the native
tetrameric enzyme, and (b) ATP is probably the major metabolic effector of
germinating castor endosperm cytosolic pyruvate kinase in vivo.
