Project description:Intact mass analyses (HPLC-ESI-MS) of GELFREE separated fractions of yeast. A total of 66 raw files, representing two replicate analyses.
Project description:Proteomics is presently dominated by the "bottom-up" strategy, in which proteins are enzymatically digested into peptides for mass spectrometric identification. Although this approach is highly effective at identifying large numbers of proteins present in complex samples, the digestion into peptides renders it impossible to identify the proteoforms from which they were derived. We present here a powerful new strategy for the identification of proteoforms and the elucidation of proteoform families (groups of related proteoforms) from the experimental determination of the accurate proteoform mass and number of lysine residues contained. Accurate proteoform masses are determined by standard LC-MS analysis of undigested protein mixtures in an Orbitrap mass spectrometer, and the lysine count is determined using the NeuCode isotopic tagging method. We demonstrate the approach in analysis of the yeast proteome, revealing 8637 unique proteoforms and 1178 proteoform families. The elucidation of proteoforms and proteoform families afforded here provides an unprecedented new perspective upon proteome complexity and dynamics.
Project description:The “quality” rather than “quantity” of lipoprotein particles is more relevant to cardiovascular diseases (CVD), since lipoprotein particles are highly heterogeneous. However, the targeted separation, detection, and exploration of the heterogeneous on omic-scale for lipoprotein remain challenging and rarely touched. Here, we established a high-resolution proteomics and lipidomics (HiPL) method. Briefly, the lipoprotein particles were separated into nine fractions (Lipo-HiPL) using anion-exchange chromatography (AEC), in which HDL, LDL, and VLDL were fractionated into three more fractions respectively. Furthermore, the HDL subclasses were delicately purified within nine fractions (HDL-HiPL), covering diverse protein-lipid heterogeneity. Integration of proteomic and lipidomic analysis revealed proteome-lipidome connectivity (PLC) in lipoprotein and HDL particles, which highly correlated with the atherosclerotic phenotype in rabbit model. Importantly, the lipoprotein “quality” revealed by PLC was found to be related more closely to clinical status in acute coronary syndrome patients than direct proteome and lipidome quantification. Thus, proteome-lipidome connectivity (PLC) as novel HDL “quality” features could be highly associated with atherosclerotic cardiovascular diseases.
Project description:This data set contains the following:
1) 66 raw files for NeuCode-labeled human proteoforms from the Jurkat cell line analyzed via "intact-mass" (i.e., LC-MS, with no precursor fragmentation). Three biological replicates were performed (files from the different replicates are labeled with 022317, 031617, or 031817, respectively). For each replicate, proteoforms were separated offline using a 12% Tris-acetate Gelfree cartridge and 11 fractions were collected. Two technical replicate LC-MS injections of each fraction were performed, yielding a total of 66 raw files (3 biological replicates x 11 fractions x 2 injections).
2) 22 raw files for label-free human proteoforms from the Jurkat cell line analyzed via "top-down" (i.e., LC-MS/MS, with precursor fragmentation). One biological replicate was performed (labeled 032017). Proteoforms were separated offline using a 12% Tris-acetate Gelfree cartridge and 11 fractions were collected. Two technical replicate LC-MS/MS injections of each fraction were performed, yielding a total of 22 raw files (1 biological replicate x 11 fractions x 2 injections).
3) Multi-protease and trypsin-only pruned G-PTM-D databases used for proteoform identification. Note that the bottom-up data used to generate these databases can be found elsewhere on MassIVE (MSV000083304).
Project description:Raw top-down LC-MS/MS files from 12% GELFREE separated fractions of yeast. Raw files for 2 technical replicates of each fraction, in total 24 raw files.