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Core histones and their PTMs play important roles in regulating gene transcription and other DNA-related processes. The study of core histones PTMs, their cross-talk and functional roles is not only of broad biological significance but also of wide pathological and clinical relevance. Having the strength of comprehensive proteoform identification with 100% amino acid sequence coverage and combinatorial PTMs, top-down proteomics has become the state-of-the-art analytical tool for combinatorial PTM characterization of core histones. In this study, we report our top-down characterization of chicken (Gallus gallus domesticus) core histones, which have been widely used as models for chromosome re-construction among others because of easy availability and not-so-dense PTMs. With nanoRPLC-MS/MS analysis and ProteinGoggle database search, a total of 58 proteoforms were identified for the core histone families of H4, H2B, H2A, and H3. [ABSTRACT FROM AUTHOR]

For top-down protein database search and identification from tandem mass spectra, our isotopic envelope fingerprinting search algorithm and ProteinGoggle search engine have demonstrated their strength of efficiently resolving heavily overlapping data as well separating non-ideal data with non-ideal isotopic envelopes from ideal ones with ideal isotopic envelopes. Here we report our updated ProteinGoggle 2.0 for intact protein database search with full-capacity. The indispensable updates include users' optional definition of dynamic post-translational modifications and static chemical labeling during database creation, comprehensive dissociation methods and ion series, as well as a Proteoform Score for each proteoform. ProteinGoggle has previously been benchmarked with both collision-based dissociation (CID, HCD) and electron-based dissociation (ETD) data of either intact proteins or intact proteomes. Here we report our further benchmarking of the new version of ProteinGoggle with publically available photon-based dissociation (UVPD) data ( http://hdl.handle.net/2022/17316 ) of intact E. coli ribosomal proteins. Biological significance Protein species (aka proteoforms) function at their molecular level, and diverse structures and biological roles of every proteoform come from often co-occurring proteolysis, amino acid variation and post-translational modifications. Complete and high-throughput capture of this combinatorial information of proteoforms has become possible in evolving top-down proteomics; yet, various methods and technologies, especially database search and bioinformatics identification tools, in the top-down pipeline are still in their infancy stages and demand intensive research and development. [ABSTRACT FROM AUTHOR]