Substrate kinetics and substrate effects on the quaternary structure of barley UDP-glucose pyrophosphorylase.
2012 (English)In: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 79, 39-45 p.Article in journal (Refereed) Published
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.
Place, publisher, year, edition, pages
2012. Vol. 79, 39-45 p.
Biochemistry and Molecular Biology
IdentifiersURN: urn:nbn:se:umu:diva-58610DOI: 10.1016/j.phytochem.2012.04.002ISI: 000307031800003PubMedID: 22552276OAI: oai:DiVA.org:umu-58610DiVA: diva2:549270