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. 2016 Jul 12;44(20):9942–9955. doi: 10.1093/nar/gkw631

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© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 1. — TRBP-D3 is a substrate of S6 kinases. (A) Alignment of candidate S6K phosphorylation sites on TRBP and known sites on S6RP (shown in green). A model S6K substrate peptide is also shown for reference. Two validated S6K substrates containing the RXRXXS or RXXS consensus motifs (R in purple) are also shown. (B) TRBP-D3 (aa258-366) and PACT-D3 (aa208–313) sequence alignment, illustrating known TRBP and PACT phosphorylation sites (green). (C) [γ-32P] autoradiogram and total protein (detected by Coomassie staining) after kinase assay incubation (4 h) of TRBP-D3 and TRBP-D3(DD) with active S6K1, S6K2, or AKT1. (D) [γ-32P] autoradiogram and total protein (detected by Coomassie staining) after kinase assay incubation (4 h) of TRBP-D1/D2 with recombinant S6K1, S6K2, AKT1 or ERK2. A band shift due to ERK phosphorylation is observed for TRBP-D1/D2. (E) [γ-32P] counts for TRBP-D3 and PACT-D3 during 4 h in vitro phosphorylation by recombinant S6K2.