Q exactive hf ultra high mass range qe uhmr

Nucleosomes samples at a concentration of 2 μM and desalted into 150 mM ammonium acetate using 30 kDa MWCO 0.5 mL spin filters (Millipore-Sigma). Samples were analyzed using a Q Exactive HF mass spectrometer with Extended Mass Range (QE-EMR) and a Q Exactive HF Ultra-High Mass Range (QE-UHMR), both by Thermo Fisher Scientific. Data were collected using XCalibur QualBrowser 4.0.27.10 (Thermo Fisher Scientific). The native electrospray platform is coupled to a three-tiered tandem MS process. First, the analysis of the intact nucleosome (MS¹) provides the total complex mass (reported as a deconvoluted neutral average mass value).^{13 (link)} In stage two, the nucleosome is activated by collisions with nitrogen gas to eject histones (MS²). In stage three, further vibrational activation of the ejected histones via collisions with nitrogen gas yields backbone fragmentation products from each monomer (MS³) that are recorded at isotopic resolution (120,000 resolving power at m/z 400). These fragments can be mapped onto the primary sequence of the histones in order to localize posttranslational modifications. A step-by-step protocol for Nuc-MS data acquisition and analysis is available on the Nature Protocol Exchange repository (DOI 10.21203/rs.3.pex-1288/v1) and in the Supplemental Information.³⁴