SEC-MS of the HSP90 inhibitor-modulated proteome in HT29 human colon cancer cells

HT29 human colon adenocarcinoma cells treated with the HSP90 inhibitor tanespimycin/17-AAG ('HSP90i') or DMSO vehicle ('Control') were lysed under non-denaturing conditions and fractionated by size-exclusion chromatography (SEC) to separate native protein complexes according to their molecular weight (fraction 1 = largest). Each of the 48 fractions (24 per treatment condition) were subjected to mass spectrometry (MS)-based bottom-up proteomics. 4,645 proteins were quantified in one or more fractions in at least 3 of the 4 Experiments (biological replicates) for Control or HSP90i conditions. Summed fraction intensities (i.e., the sum of Label-Free Quantitation (LFQ) intensities across all 24 fractions in each experiment) were used for differential expression analysis, depicted in the searchable table and Volcano Plot. Tukey plots of summed intensities, and SEC-MS profiles, will be plotted for up to 8 proteins selected from the table below. For SEC-MS profiles: lines and ribbons represent mean LFQ intensities and standard errors, respectively, across Experiments. The green vertical line represents estimated fraction of the monomer, based on molecular weight.

For more information, please see publication Native size exclusion chromatography-based mass spectrometry (SEC-MS) identifies novel components of the Heat Shock Protein 90-dependent proteome', by Samant, Batista, et al. (2022).

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