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. 2018 Apr 12;8:5903. doi: 10.1038/s41598-018-24068-w

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Ssa1/Hsp70 binds Aco1 and its C and N terminal domains. (A) Identification of proteins associated with Aco1-C by LC-MS/MS. GST-C or GST-CΔ6, expressed in yeast, were pulled down from lysates using glutathione beads. Proteins were released from the beads and digested with trypsin in solution or run on PAGE followed by in-gel digestion with trypsin. The peptides associated with the Aco1 C-terminus were identified by LC-MS/MS. The number of peptides and coverage of the protein sequence in each case is presented. (B) Full length aconitase (GST-Aco1), GST-Aco1-C and GST-Aco1-N can pull-down Ssa1 from yeast lysates. Lysates of yeast cells expressing the indicated fusion proteins were incubated with glutathione sepharose beads, which were subsequently washed and resuspended in SDS containing loading buffer. Samples were subjected to Western blot analysis using anti-Ssa1 antiserum (GenScript. Lot number-642114S01/P20011507). (C) Structural model of the Aco1 - Ssa1 complex. The predicted complex structure was created by docking the structural models of Ssa1 and Aco1 (generated using I-Tasser) using PatchDock and then refining it using RosettaDock. Aco1-N, light green; Aco1-C, light blue; Ssa1, purple.