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BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES

Heat Stability and Allosteric Properties of the Maize Endosperm ADP-Glucose Pyrophosphorylase Are Intimately Intertwined^1,[OA]

Program in Plant Molecular and Cellular Biology and Horticultural Sciences (S.K.B., J.R.S., L.C.H.), and Department of Chemistry (J.D.S.), University of Florida, Gainesville, Florida 32611–7200

ADP-glucose (Glc) pyrophosphorylase (AGPase), a key regulatory enzyme in starch biosynthesis, is highly regulated. Transgenic approaches in four plant species showed that alterations in either thermal stability or allosteric modulation increase starch synthesis. Here, we show that the classic regulators 3-phosphoglyceric acid (3-PGA) and inorganic phosphate (Pi) stabilize maize (Zea mays) endosperm AGPase to thermal inactivation. In addition, we show that glycerol phosphate and ribose-5-P increase the catalytic activity of maize AGPase to the same extent as the activator 3-PGA, albeit with higher K_a (activation constant) values. Activation by fructose-6-P and Glc-6-P is comparable to that of 3-PGA. The reactants ATP and ADP-Glc, but not Glc-1-P and pyrophosphate, protect AGPase from thermal inactivation, a result consistent with the ordered kinetic mechanism reported for other AGPases. 3-PGA acts synergistically with both ATP and ADP-Glc in heat protection, decreasing the substrate concentration needed for protection and increasing the extent of protection. Characterization of a series of activators and inhibitors suggests that they all bind at the same site or at mutually exclusive sites. Pi, the classic "inhibitor" of AGPase, binds to the enzyme in the absence of other metabolites, as determined by thermal protections experiments, but does not inhibit activity. Rather, Pi acts by displacing bound activators and returning the enzyme to its activity in their absence. Finally, we show from thermal inactivation studies that the enzyme exists in two forms that have significantly different stabilities and do not interconvert rapidly.

The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: L. Curtis Hannah (hannah{at}mail.ifas.ufl.edu).

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www.plantphysiol.org/cgi/doi/10.1104/pp.107.109942

^* Corresponding author; e-mail address hannah{at}mail.ifas.ufl.edu.

Received September 27, 2007; accepted November 7, 2007; published November 16, 2007.

This article has been cited by other articles:

				S. K. Boehlein, J. R. Shaw, J. D. Stewart, and L. C. Hannah Characterization of an Autonomously Activated Plant ADP-Glucose Pyrophosphorylase Plant Physiology, January 1, 2009; 149(1): 318 - 326. [Abstract] [Full Text] [PDF]