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. 2016 Feb 29;5(2):e201. doi: 10.1038/oncsis.2016.14

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Copyright © 2016 Macmillan Publishers Limited

Oncogenesis is an open-access journal published by Nature Publishing Group. This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

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Identification of HSP60 as the target of P17. (a) Protocols used for pull-down-assay-based analysis of binder of P17. (b) Biotin-P17 labeled streptavidin magnetic beads were used for pull-down assay from TRAIL-treated Jurkat cell extracts. Bound proteins were separated by SDS–PAGE. Absent from TRAIL-untreated and bead-control samples, a major band at 58 kDa was excised for mass spectrometry and identified as HSP60. (c) Mass spectrometry analysis identified peptides in HSP60 with 37% sequence coverage. Matched amino acids were shown in red. (d) Colocalization of FITC-P17 and anti-HSP60 (Alexa Fluor 594-conjugated) in TRAIL-treated Jurkat cells (Scale bars: 10 μm) observed by confocal microscopy (upper) and analyzed by Image-Pro Plus (bottom). (e) The schematic illustration for pull-down and western blot analysis. (f) Detection of HSP60 in pull-down samples by western blot and coomassie brilliant blue staining.