Improved A. tumefaciens Glycogen Synthase by addition of an SBD from A. thaliana Starch Synthase III

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

Mendoza

Congreso; XLVIII Reunion SAIB; 2012

SAIB

Starch synthase III (SSIII) from Arabidopsis thaliana is a SS isoform with a particular primary structure organization; the C-terminal domain, highly conserved in other SS isoforms, is preceded by a unique specific domain (SSIII-SD) which contains three in tandem starch binding domains (SBDs, named D1, D2 and D3) characteristic of degrading enzymes. These N-terminal SBDs have a probed regulatory role in SSIII activity, showing starch binding ability and modulating the catalytic properties of the enzyme. To further investigate the functional role of A. thaliana SSIII-SD, three His tagged chimeric proteins were constructed combining the SBDs from A. thaliana with the glycogen synthase (GS) from Agrobacterium tumefaciens, which lacks SBDs. Recombinant quimeric proteins were expressed and purified to homogeneity from Escherichia coli in order to kinetically characterize them. Furthermore, quimeric enzymes capability to restore in vivo glycogen biosynthesis in E. coli cells was also tested. The results obtained showed that the D3-GS enzyme showed increased capacity of glycogen synthesis in vivo with minor changes in their in vitro kinetics parameters.   Resumen: 1406 caracteres