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

Decker, DanielUmeå universitet,Institutionen för fysiologisk botanik,Umeå Plant Science Centre (UPSC) (författare)

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.

NATURVETENSKAP Biologi Biokemi och molekylärbiologi hsv//swe
NATURAL SCIENCES Biological Sciences Biochemistry and Molecular Biology hsv//eng
Cell wall synthesis
Oligomerization
Protein structure
Sucrose metabolism
Sucrose synthase
Sugar activation
UDP-sugar synthesis

Meng, MengUmeå universitet,Institutionen för fysiologisk botanik,Umeå Plant Science Centre (UPSC)(Swepub:umu)meme0001 (författare)
Gornicka, AgnieszkaUmeå universitet,Institutionen för fysiologisk botanik,Umeå Plant Science Centre (UPSC) (författare)
Hofer, AndersUmeå universitet,Institutionen för medicinsk kemi och biofysik(Swepub:umu)anho0002 (författare)
Wilczynska, MalgorzataUmeå universitet,Institutionen för medicinsk kemi och biofysik(Swepub:umu)mawi0006 (författare)
Kleczkowski, Leszek AUmeå universitet,Institutionen för fysiologisk botanik,Umeå Plant Science Centre (UPSC)(Swepub:umu)lekl0001 (författare)
Umeå universitetInstitutionen för fysiologisk botanik (creator_code:org_t)

Av författaren/redakt...: Decker, Daniel; Meng, Meng; Gornicka, Agnies ...; Hofer, Anders; Wilczynska, Malg ...; Kleczkowski, Les ...

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