Laguillo-Gómez, AndreaCalvo, EnriqueMartín-Cófreces, NoaLozano-Prieto, MartaSánchez-Madrid, FranciscoVazquez, Jesus2023-10-172023-10-172023-09-15J Proteomics. 2023 Sep 15:287:104968.http://hdl.handle.net/20.500.12105/16576Open-search methods allow unbiased, high-throughput identification of post-translational modifications in proteins at an unprecedented scale. The performance of current open-search algorithms is diminished by experimental errors in the determination of the precursor peptide mass. In this work we propose a semi-supervised open search approach, called ReCom, that minimizes this effect by taking advantage of a priori known information from a reference database, such as Unimod or a database provided by the user. We present a proof-of-concept study using Comet-ReCom, an improved version of Comet-PTM. Comet-ReCom increased identification performance of Comet-PTM by 68%. This increased performance of Comet-ReCom to score the MS/MS spectrum comes in parallel with a significantly better assignation of the monoisotopic peak of the precursor peptide in the MS spectrum, even in cases of peptide coelution. Our data demonstrate that open searches using ultra-tolerant mass windows can benefit from using a semi-supervised approach that takes advantage from previous knowledge on the nature of protein modifications. SIGNIFICANCE: The present study introduces a novel approach to ultra-tolerant database search, which employs prior knowledge of post-translational modifications (PTMs) to improve identification of modified peptides. This method addresses the limitations related to experimental errors and precursor mass assignation of previous open-search methods. Thus, it enables the study of the biological significance of a wider variety of PTMs, including unknown or unexpected modifications that may have gone unnoticed using non-supervised search methods.engVoRhttp://creativecommons.org/licenses/by-nc-nd/4.0/Tandem Mass SpectrometryPeptidesAmino Acid SequenceProteinsAlgorithmsProtein Processing, Post-TranslationalDatabases, ProteinSoftwareAttribution-NonCommercial-NoDerivatives 4.0 Internacional3746362228710496810.1016/j.jprot.2023.1049681876-7737Journal of proteomicsopen access