Identification and Characterization of a Critical Region in the Glycogen Synthase from Escherichia coli

The cysteine-specific reagent 5,5′-dithiobis(2-nitrobenzoic acid) inactivates the Escherichia coli glycogen synthase (Holmes, E., and Preiss, J. (1982) Arch. Biochem. Biophys. 216, 736-740). To find the responsible residue, all cysteines, Cys7, Cys379, and Cys408, were substituted combinatorially by Ser. 5,5′-Dithiobis(2-nitrobenzoic acid) modified and inactivated the enzyme if and only if Cys379 was present and it was prevented by the substrate ADP-glucose (ADP-Glc). Mutations C379S and C379A increased the S0.5 for ADP-Glc 40- and 77-fold, whereas the specific activity was decreased 5.8- and 4.3-fold, respectively. Studies by and Cys379 was the the enzyme the and for Cys379 were by by and decreased the for ADP-Glc and glycogen by decreased only the for The activity the enzyme was by the and were by the the The for and substrate The cysteine-specific reagent 5,5′-dithiobis(2-nitrobenzoic acid) inactivates the Escherichia coli glycogen synthase (Holmes, E., and Preiss, J. (1982) Arch. Biochem. Biophys. 216, 736-740). To find the responsible residue, all cysteines, Cys7, Cys379, and Cys408, were substituted combinatorially by Ser. 5,5′-Dithiobis(2-nitrobenzoic acid) modified and inactivated the enzyme if and only if Cys379 was present and it was prevented by the substrate ADP-glucose (ADP-Glc). Mutations C379S and C379A increased the S0.5 for ADP-Glc 40- and 77-fold, whereas the specific activity was decreased 5.8- and 4.3-fold, respectively. Studies by and Cys379 was the the enzyme the and for Cys379 were by by and decreased the for ADP-Glc and glycogen by decreased only the for The activity the enzyme was by the and were by the the The for and substrate Escherichia coli glycogen synthase the ADP-glucose glycogen glycogen the was J. J. and the J. J. The glycogen synthase coli J. J. and J. Arch. Biochem. Biophys. J. J. J. J. J. glycogen and the and and and ADP-Glc and whereas the and glycogen and the and by and by J. J. Biochem. glycogen synthase and glycogen J. 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