Dibavar, A. S. | Search bioRxiv

bioRxiv2022

Monitoring Functional Post-Translational Modifications Using a Data-Driven Proteome Informatic Pipeline Based on PEIMAN2

Post-translational modifications (PTMs) are of significant interest in molecular biomedicine due to their crucial role in signal transduction across various cellular and organismal processes. Characterizing PTMs, distinguishing between functional and inert modifications, quantifying their occupancies, and understanding PTM crosstalk are challenging tasks in any biosystem. Studying each PTM often requires a specific, labor- intensive experimental design. Here, we present a PTM-centric proteome informatic pipeline for predicting relevant PTMs in mass spectrometry-based proteomics data without prior information. Once predicted, these in silico identified PTMs can be incorporated into a refined database search and compared to measured data. As a practical application, we demonstrate how this pipeline can be used to study glycoproteomics in oral squamous cell carcinoma based on the proteome profile of primary tumors. Subsequently, we experimentally identified cellular proteins that are differentially expressed in cells treated with multikinase inhibitors dasatinib and staurosporine using mass spectrometry-based proteomics. Computational enrichment analysis was then employed to determine the potential PTMs of differentially expressed proteins induced by both drugs. Finally, we conducted an additional round of database search with the predicted PTMs. Our pipeline successfully analyzed the enriched PTMs, and detected proteins not identified in the initial search. Our findings support the effectiveness of PTM-centric searching of MS data in proteomics based on computational enrichment analysis, and we propose integrating this approach into future proteomics search engines.

biochemistry↗

bioRxiv2021

Abnormal (hydroxy)prolines deuterium content redefines hydrogen chemical mass

Analysing the {delta}2H in individual amino acids of proteins extracted from vertebrates, we unexpectedly found in some samples, notably bone collagen from seals, more than twice as much deuterium in proline and hydroxyproline residues than in seawater. This corresponds to at least four times higher {delta}2H than in any previously reported biogenic sample. We ruled out diet as a plausible mechanism for such anomalous enrichment. This finding puts into question the old adage that you are what you eat. SUMMARYThe chemical mass of hydrogen is defined as an interval from the lowest to the highest content of deuterium 2H, hydrogens heavy stable isotope. Measurements of the deviations {delta}2H in the deuterium content from the standard (ocean water, {delta}2H = 0{per thousand}) are used to characterise biological samples, such as animal bone collagen. The results are often interpreted in terms of the trophic level and diet of the animal as well as prevailing climate during its lifetime. The majority of the published bone collagen {delta}2H data fall into a narrow {delta}2H range limited to {+/-}100{per thousand}. Using novel analysis method, we unexpectedly found greatly higher {delta}2H values, up to 1500{per thousand}, in seal bone collagen. Such anomalous deuterium enrichment is detected only in two amino acid residues, proline and its derivative hydroxyproline, while other residues show much smaller {delta}2H values. Anomalously high {delta}2H values, albeit of lower magnitudes, are also found for these residues in other biological sources. This finding substantially expands the upper bound of the hydrogen chemical mass for biogenic sources. Since neither diet nor environment explain these mysteriously high enrichment levels amounting to more than twice deuterium content in sea water, our understanding of stable isotopes in nature, as well as the old adage "you are what you eat", are put in question.

biochemistry↗