Figure 4

Characterization of enzymatic function of recombinant Ro GAs. (A) The extracellular activity of various recombinant GAs was tested. Lane 1, pS1 (empty vector); lane 2, full-length GA; lane 3, GAΔ26–167; lane 4, GAΔ132–167; lane 5, T165A and lane 6, N167D. (B) Western blot analysis of wild-type and mutant GAs grown at 30°C (upper panel) and 20°C (lower panel) for 3 days. Lane 1, pS1 (empty vector); lane 2, full-length GA; lane 3, GAΔ26–167; lane 4, GAΔ132–167; lane 5, T165A and lane 6, N167D. (C) Starch plate assays of mutant GAs. No. 1, S. cerevisiae MNN10 cells transformed with pS1 (empty vector); No. 2, pS1-full-length GA; No. 3, pS1-GAΔ26–167; No. 4, pS1-GAΔ132–167; No. 5, pS1-T165A and No. 6, pS1-N167D. (D) The electrophoretic mobilities of wild-type GA (lane 1) and N167D (lane 2) on an 8% SDS-PAGE.