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Figure 5


Fig. 5. Glycosylation of p40 and PTP A in microsporidia. Cells from cricket fat bodies were infected with P. grylli and prepared for PAGE. (A) SDS-PAGE analysis followed by Coomassie Brilliant Blue (left) and Schiff reagent (right) staining. Lane 1, MW markers; lane 2, intact spores broken up in phosphate-buffered saline and boiled for 10 minutes in SDS-PAGE sample buffer; lane 3, intact spores boiled as for lane 2; lane 4, intact P. grylli spores in alkaline saline solution (10 mM KOH, 170 mM KCl; 30 minutes) selectively solubilizes the major 40-kDa protein; lane 5, spore extrusion followed by washing in 3% SDS and PTPs solubilized with 50% 2-mercaptoethanol; lane 6, soluble; and lane 7, membrane, proteins of the host (cricket Gryllus bimaculatus) fat body. (B) Precipitation of p40 by GNA-agarose. mw, molecular weight markers; lanes 1, 3, 5, supernatant after p40 solubilization in 10 mM KOH, 170 mM KCl; lanes 2, 4, 6, proteins precipitated by GNA-agarose and eluted by boiling in SDS-PAGE sample buffer; lanes 1, 2, pretreatment with reaction buffer for mannosidase (control); lanes 3, 4, treatment with beta-mannosidase; lanes 5, 6, treatment with {alpha}-mannosidase.





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