Profiling Changes in Histone Post-translational Modifications by Top-Down Mass Spectrometry.
Top-down mass spectrometry is a valuable tool for understanding gene expression through characterization of combinatorial histone post-translational modifications (i.e., histone code). In this protocol, we describe a top-down workflow that employs liquid chromatography (LC) coupled to mass spectrometry (MS), for fast global profiling of changes in histone proteoforms, and apply LCMS top-down approach for comparative analysis of a wild-type and a mutant fungal species. The proteoforms exhibiting differential abundances can be subjected to further targeted studies by other MS or orthogonal (e.g., biochemical) assays. This method can be generally adapted for screening of changes in histone modifications between samples such as wild type vs. mutant or healthy vs. diseased.
Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.
Mowei
M
Zhou
Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.|Department of Chemistry and Biochemistry, University of Oklahoma, Norman, OK, USA.
Si
S
Wu
Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.
David L
DL
Stenoien
Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.
Zhaorui
Z
Zhang
Department of Biochemistry and Biophysics, Center for Genome Research and Biocomputing, Oregon State University, Corvallis, OR, USA.
Lanelle
L
Connolly
Department of Biochemistry and Biophysics, Center for Genome Research and Biocomputing, Oregon State University, Corvallis, OR, USA.
Michael
M
Freitag
Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA. ljiljana.pasatolic@pnnl.gov.
Ljiljana
L
Paša-Tolić
D002852:Chromatography, Ion Exchange; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005670:Fusarium / Q000378:metabolism; D006367:HeLa Cells; D042421:Histone Code; D006657:Histones / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D011499:Protein Processing, Post-Translational; D053719:Tandem Mass Spectrometry
D005656:Fungal Proteins; D006657:Histones
Histone; Liquid chromatography; Mass spectrometry; Post-translational modification; Screening; Top-down