Figure 6.

Small RNA are the principal nucleotide substrates of the QTRT enzyme. (A) In vitro transcribed tyrosyl tRNA, (B) synthetic aspartyl stem loop, and (C) synthetic arginyl step loop (all at 3.33 μM) or (D) 1.5 μg small RNA (<200 nucleotides) or 15 μg large RNA (>200 nucleotides) isolated from MDA-MB-231 cells, were incubated with 200 nM of [³H] queuine and QTRT enzyme (175 nM) in a final reaction volume of 150 μl for 90 min at 37°C. Reaction products were separated on a DEAE cellulose column and the eluted RNA evaluated for [³H] queuine incorporation by scintillation counting. Heat-inactivated QTRT enzyme complex served as negative control. Data are mean ± SD (n=3). Representative of two independent experiments.