Ambergen Technology
tRNA Mediated Protein Engineering (TRAMPE)
Figure 3: Comparison of PC-SNAG Yield and Purity to Polyhistidine Tag Mediated Isolation
GST containing a C-terminal polyhistidine (His6) tag was cell-free expressed and TRAMPE labeled using PC-biotin and BODIPY-FL misaminoacylated tRNAs together. PC-SNAG was achieved based on the directly incorporated PC-biotins. The expression reaction mixture was split and GST was isolated either by PC-SNAG or by the His6 tag using a cobalt metal chelate affinity resin using standard procedures. (Top Panel) BODIPY-FL fluorescence SDS-PAGE image. (Bottom Panel) Silver stain for total protein of the same gel. Lanes: SB = SDS-PAGE gel loading sample buffer; -D = Isolated material derived from a blank expression reaction performed lacking only the added GST DNA; +D = Isolated material derived from an expression reaction performed with the added GST DNA. The asterisk denotes a general contaminant not attributable to the purification methods.