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. 2013 Aug 22;41(21):9825–9838. doi: 10.1093/nar/gkt741

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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 4. — Mis-charging of [³²P]CatRNA^Ser with Nva and post-transfer editing of Nva-[³²P] CatRNA^Ser by CaLeuRS and CaLeuRS-D422A. (A) A representative graph showing mis-charging of [³²P]CatRNA^Ser with non-cognate Nva. Free [³²P]CatRNA^Ser and mis-charged [³²P]CatRNA^Ser are represented by [³²P]AMP and Nva-[³²P]AMP after digestion of nuclease S1. Known amounts of [α-³²P]ATP were serially diluted and loaded onto the TLC plate after separation for quantification. (B) Quantitative analysis of Nva-[³²P] CatRNA^Ser generated by CaLeuRS (black square) and CaLeuRS-D422A (black up-pointing triangle) in (A). (C) A representative graph showing hydrolysis of Nva-[³²P]CatRNA^Ser by CaLeuRS and CaLeuRS-D422A. A control reaction represented the spontaneous hydrolysis of Nva-[³²P]CatRNA^Ser without the addition of enzyme. (D) Analysis of post-transfer editing of Nva-[³²P]CatRNA^Ser by CaLeuRS (black square) and CaLeuRS-D422A (black up-pointing triangle) in (C).