Kinetic characterization of A. tumefaciens glycogen synthase.

WAYLLACE, N. Z.; VALDEZ, H.; BUSI, M. V.; DIEGO FABIAN GOMEZ CASATI; UGALDE, R.

Rosario

Congreso; XLII Reunión SAIB; 2006

The glycogen and starch are the most widespread energy storage compounds in living organisms. Glycogen and starch synthases (GS or SSs) are glycosyltransferases that catalyses the transfer of glucosyl residues from ADPGlc to the non-reducing end of a growing alpha-1,4-glucan chain. The structural, kinetic and regulatory properties of GSs from mammals or SSs from plants were well characterized. Furthermore, it has been recently described that the 3D structure of A. tumefaciens GS posseses a fold common to other glycosyltransferases. However, the regulatory properties from bacterial GSs were less studied. In the present work, recombinant His-tagged A. tumefaciens GS was expressed in E. coli BL21 RIL cells and purified to homogeneity by one-step purification using a Ni-chelating colum. We determined that A. tumefaciens GS is active in the presence of different glucosyl donors. Indeed, the enzyme is inhibited by malate, succinate and citrate, three metabolites from the TCA cycle. These results allow us to propose the different residues that determine glucosyl donor specificity and suggest that the GS activity is determined by the levels of different compounds that participates in energetic metabolism.