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. 2013 Nov 26;42(4):2602–2623. doi: 10.1093/nar/gkt1228

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© The Author(s) 2013. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 3. — In vitro and in vivo activity of the recombinant proteins MnmC(o) and MnmC(m). (A) and (B) HPLC analysis of total tRNA from a null mnmC mutant (IC6019; panel A) and an mnmC(m)-G68D mutant (IC6018; panel B) before (solid black line) and after in vitro incubation (dotted red line) with purified MnmC(o) and MnmC(m) recombinant proteins, respectively. (C) and (D) HPLC analysis of total tRNA extracted from IC6019 (panel C) and IC6018 (panel D) transformed with pBAD-TOPO (solid black lines) or a pBAD-TOPO derivative expressing Flag-MnmC(o) and Flag-MnmC(m), respectively (dotted red lines). Absorbance was monitored at 314 nm to maximize the detection of thiolated nucleosides. Small variations in elution times between upper and lower panels were probably due to negligible variations in buffers prepared in different days. Note that nucleosides were identified by both elution times and spectra (data not shown).