{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,31]],"date-time":"2026-03-31T16:01:31Z","timestamp":1774972891771,"version":"3.50.1"},"reference-count":65,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2021,4,30]],"date-time":"2021-04-30T00:00:00Z","timestamp":1619740800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100008530","name":"European Regional Development Fund","doi-asserted-by":"publisher","award":["IU16-015983"],"award-info":[{"award-number":["IU16-015983"]}],"id":[{"id":"10.13039\/501100008530","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004587","name":"Instituto de Salud Carlos III","doi-asserted-by":"publisher","award":["PI16\/00346 and PI20\/00936"],"award-info":[{"award-number":["PI16\/00346 and PI20\/00936"]}],"id":[{"id":"10.13039\/501100004587","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004587","name":"Instituto de Salud Carlos III","doi-asserted-by":"publisher","award":["PT17\/0019\/0022"],"award-info":[{"award-number":["PT17\/0019\/0022"]}],"id":[{"id":"10.13039\/501100004587","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Sara Borrell Postdoctoral Fellowship, Acci\u00f3n Estrat\u00e9gica en Salud","award":["CD17\/00109"],"award-info":[{"award-number":["CD17\/00109"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Proteomes"],"abstract":"<jats:p>Protamines replace histones as the main nuclear protein in the sperm cells of many species and play a crucial role in compacting the paternal genome. Human spermatozoa contain protamine 1 (P1) and the family of protamine 2 (P2) proteins. Alterations in protamine PTMs or the P1\/P2 ratio may be associated with male infertility. Top-down proteomics enables large-scale analysis of intact proteoforms derived from alternative splicing, missense or nonsense genetic variants or PTMs. In contrast to current gold standard techniques, top-down proteomics permits a more in-depth analysis of protamine PTMs and proteoforms, thereby opening up new perspectives to unravel their impact on male fertility. We report on the analysis of two normozoospermic semen samples by top-down proteomics. We discuss the difficulties encountered with the data analysis and propose solutions as this step is one of the current bottlenecks in top-down proteomics with the bioinformatics tools currently available. Our strategy for the data analysis combines two software packages, ProSight PD (PS) and TopPIC suite (TP), with a clustering algorithm to decipher protamine proteoforms. We identified up to 32 protamine proteoforms at different levels of characterization. This in-depth analysis of the protamine proteoform landscape of normozoospermic individuals represents the first step towards the future study of sperm pathological conditions opening up the potential personalized diagnosis of male infertility.<\/jats:p>","DOI":"10.3390\/proteomes9020021","type":"journal-article","created":{"date-parts":[[2021,4,30]],"date-time":"2021-04-30T10:55:04Z","timestamp":1619780104000},"page":"21","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Protamine Characterization by Top-Down Proteomics: Boosting Proteoform Identification with DBSCAN"],"prefix":"10.3390","volume":"9","author":[{"given":"Gianluca","family":"Arauz-Garofalo","sequence":"first","affiliation":[{"name":"Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology, Baldiri Reixac, 10, 08028 Barcelona, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3272-0163","authenticated-orcid":false,"given":"Meritxell","family":"Jodar","sequence":"additional","affiliation":[{"name":"Molecular Biology of Reproduction and Development Research Group, Institut d\u2019Investigacions Biom\u00e8diques August Pi i Sunyer (IDIBAPS), Fundaci\u00f3 Cl\u00ednic per a la Recerca Biom\u00e8dica (FCRB), Faculty of Medicine and Health Sciences, University of Barcelona, 08036 Barcelona, Spain"},{"name":"Biochemistry and Molecular Genetics Service, Hospital Cl\u00ednic de Barcelona, 08036 Barcelona, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mar","family":"Vilanova","sequence":"additional","affiliation":[{"name":"Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology, Baldiri Reixac, 10, 08028 Barcelona, 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Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ada","family":"Soler-Ventura","sequence":"additional","affiliation":[{"name":"Molecular Biology of Reproduction and Development Research Group, Institut d\u2019Investigacions Biom\u00e8diques August Pi i Sunyer (IDIBAPS), Fundaci\u00f3 Cl\u00ednic per a la Recerca Biom\u00e8dica (FCRB), Faculty of Medicine and Health Sciences, University of Barcelona, 08036 Barcelona, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4876-2410","authenticated-orcid":false,"given":"Rafael","family":"Oliva","sequence":"additional","affiliation":[{"name":"Molecular Biology of Reproduction and Development Research Group, Institut d\u2019Investigacions Biom\u00e8diques August Pi i Sunyer (IDIBAPS), Fundaci\u00f3 Cl\u00ednic per a la Recerca Biom\u00e8dica (FCRB), Faculty of Medicine and Health Sciences, University of Barcelona, 08036 Barcelona, Spain"},{"name":"Biochemistry and 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Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,4,30]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1016\/S0079-6603(08)60839-9","article-title":"Vertebrate protamine genes and the histone-to-protamine replacement reaction","volume":"40","author":"Oliva","year":"1991","journal-title":"Prog. 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