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Figure 2


Fig. 2. Nuclease activity of recombinant T. brucei endoG protein ({alpha}-rTbEG). (A) Polyclonal antibodies raised against full-length T. brucei endoG protein reacted with procyclic T. brucei lysates (L) and the TbEG recombinant protein (R). Pre-immune serum was used as a control (NRS). (B) A nuclease cleavage assay was performed with wild-type (rTbEG) or mutant TbEG (H220A and E261A, rTbEGm) proteins using genomic DNA (gDNA). Treatment with aurintricarboxylic acid (ATA) inhibited DNA degradation, confirming the specificity of TbEG for nucleic acid substrates. (C) Nuclease assay as in B, using total RNA as a substrate. (D) TUNEL assay of p-formaldehyde-fixed T. brucei procyclic cells incubated with rTbEG (light black line) or rTbEGm protein (dark black line) analyzed by flow cytometry. Untreated T. brucei cells (gray shaded peak) served as a control.