The Clinical Proteomic Tumor Analysis Consortium (CPTAC) analyzes cancer biospecimens by mass spectrometry, characterizing and quantifying their constituent proteins, or proteome. Proteomic analysis for each CPTAC study is carried out independently by Proteomic Characterization Centers (PCCs) using a variety of protein fractionation techniques, instrumentation, and workflows. Mass spectrometry and...

Description from PRIDE: "The NRF2 transcription factor controls a cell stress program that is implicated in cancer and there is great interest in targeting NRF2 for therapy. We show that NRF2 activity depends on Fructosamine-3-kinase (FN3K) - a kinase that triggers protein de-glycation. In its absence, NRF2 is extensively glycated, unstable, and defective at binding to small MAF proteins and transcriptional...

Description from PRIDE: "Sex chromosomes in males of most mammalian species share only a small homologous segment, the pseudoautosomal region (PAR), wherein double-strand break (DSB) formation must occur for correct meiotic segregation. We find that multiple DSB-promoting factors hyperaccumulate in the PAR, including the ankyrin-repeat domain-containing protein ANKRD31. We used immunoprecipitation...

Spindle assembly checkpoint (SAC) regulators such as the Mps1 kinase not only delay anaphase onset, but also correct improper chromosome-spindle linkages that would otherwise lead to missegregation and aneuploidy. However, the substrates and mechanisms involved in this pathway of error correction remain poorly understood. Using a chemically tuned kinetochore-targeting assay, we show that Mps1 destabilizes...

To improve identification of canonical and non-canonical protein isoforms, we introduced ProteomeGenerator, a framework for reference-guided and de novo proteogenomic database generation from transcriptomic sequencing dataset. The proteomic databases output by ProteomeGenerator contain only proteins encoded by actively transcribed genes, and includes sample-specific protein isoforms resulting from...

Evaluation of sensitivity and accuracy of widely used scoring functions Sequest, MaxQuant, Peaks, and Byonic. Spectra from E. coli were matched to human sequences, and human spectra from K052 cells were matched to A. loki sequences to determine specificity of matching and FDR estimation. A subsampling analysis of SCX-RP fractionation was performed.

Reactive cellular metabolites can modify macromolecules and form adducts known as non-enzymatic covalent modifications (NECMs). Dissecting the mechanisms, regulation and consequences of NECMs, such as glycation, has been challenging due to the complex and often ambiguous nature of the adducts formed. Directly tracking the formation of modifications on key targets to uncover their underlying physiological...

Inflammasomes are multiprotein complexes formed in response to pathogens. NLRP1 and CARD8 are related proteins that form inflammasomes, but the pathogen-associated signal(s) and the molecular mechanisms controlling their activation have not been established. Inhibitors of the serine dipeptidyl peptidases DPP8 and DPP9 (DPP8/9) were recently discovered to activate both NLRP1 and CARD8. Interestingly,...

Recent development of mass spectrometer cleavable protein cross-linkers and algorithms for their spectral identification now permits large-scale cross-linking mass spectrometry (XL-MS). Here, we optimized the use of cleavable disuccinimidyl sulfoxide (DSSO) cross-linker for labeling native protein complexes in live human cells. We applied a generalized linear mixture model to calibrate cross-link...

Here, we introduce a simple method, termed CHOPS, for the discovery of protease substrates. CHOPS exploits a 2-pyridinecarboxaldehyde (2PCA)-biotin probe, which selectively biotinylates protein N-termini except those with proline in the second position. CHOPS can, in theory, discover substrates for any protease, but is particularly well-suited to discover canonical DPP substrates, as cleaved but not...