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Phytochemistry. 2012 Jul;79:39-45. doi: 10.1016/j.phytochem.2012.04.002. Epub 2012 May 1.

Substrate kinetics and substrate effects on the quaternary structure of barley UDP-glucose pyrophosphorylase.

Author information

1
Department of Plant Physiology, Umeå Plant Science Centre, Umeå University, 90187 Umeå, Sweden.

Abstract

UDP-Glc pyrophosphorylase (UGPase) is an essential enzyme responsible for production of UDP-Glc, which is used in hundreds of glycosylation reactions involving addition of Glc to a variety of compounds. In this study, barley UGPase was characterized with respect to effects of its substrates on activity and quaternary structure of the protein. Its K(m) values with Glc-1-P and UTP were 0.33 and 0.25 mM, respectively. Besides using Glc-1-P as a substrate, the enzyme had also considerable activity with Gal-1-P; however, the K(m) for Gal-1-P was very high (>10 mM), rendering this reaction unlikely under physiological conditions. UGPase had a relatively broad pH optimum of 6.5-8.5, regardless of the direction of reaction. The enzyme equilibrium constant was 0.4, suggesting slight preference for the Glc-1-P synthesis direction of the reaction. The quaternary structure of the enzyme, studied by Gas-phase Electrophoretic Mobility Macromolecule Analysis (GEMMA), was affected by addition of either single or both substrates in either direction of the reaction, resulting in a shift from UGPase dimers toward monomers, the active form of the enzyme. The substrate-induced changes in quaternary structure of the enzyme may have a regulatory role to assure maximal activity. Kinetics and factors affecting the oligomerization status of UGPase are discussed.

PMID:
22552276
DOI:
10.1016/j.phytochem.2012.04.002
[Indexed for MEDLINE]

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