Proteomes最新文献

{"title":"Neurogranin as a Synaptic Biomarker in Mild Traumatic Brain Injury: A Systematic Review of Diagnostic and Pathophysiological Evidence.","authors":"Ioannis Mavroudis, Foivos Petridis, Eleni Karantali, Dimitrios Kazis","doi":"10.3390/proteomes13030046","DOIUrl":"10.3390/proteomes13030046","url":null,"abstract":"<p><p>Neurogranin (NRGN), a synaptic protein essential for plasticity and memory function, is gaining recognition as a promising biomarker for mild traumatic brain injury (mTBI). This systematic review brings together findings from six studies that measured neurogranin levels in biofluids-including serum, cerebrospinal fluid (CSF), plasma, and exosomes-during both the acute and chronic phases following injury. In the acute phase of mTBI, elevated levels of neurogranin were consistently observed in serum samples, suggesting its potential as a diagnostic marker. These increases appear to reflect immediate synaptic disturbances caused by injury. In contrast, studies focusing on the chronic phase reported a decrease in exosomal neurogranin levels, pointing to ongoing synaptic dysfunction well after the initial trauma. This temporal shift in neurogranin expression highlights its dual utility-both as an early indicator of injury and as a longer-term marker of synaptic integrity. However, interpreting these findings is not straightforward. The studies varied considerably in terms of sample type, timing of measurements, and control for potential confounding factors such as physical activity. Such variability makes direct comparisons difficult and may influence the outcomes observed. Additionally, none of the studies included proteoform-specific analyses of neurogranin, an omission that limits our understanding of the molecular changes underlying mTBI-related synaptic alterations. Due to heterogeneity across study designs and outcome measures, a meta-analysis could not be performed. Instead, a narrative synthesis was conducted, revealing consistent patterns in neurogranin dynamics over time and underscoring the influence of biofluid selection on measured outcomes. Overall, the current evidence supports neurogranin's potential as both a diagnostic and mechanistic biomarker for mTBI. Yet, to fully realize its clinical utility, future research must prioritize standardized protocols, the inclusion of proteoform profiling, and rigorous longitudinal validation studies.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452557/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113808","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proteomic Analysis of Mechanical Injury Effects in Papaya Fruit at Two Maturity Stages.","authors":"Francisco Antonio Reyes-Soria, Eliel Ruiz-May, Enrique Castaño, Miguel Ángel Herrera-Alamillo, José Miguel Elizalde-Contreras, Samuel David Gamboa-Tuz, Lidia F E Huerta-Nuñez, Jesús Alejandro Zamora-Briseño, Luis Carlos Rodríguez-Zapata","doi":"10.3390/proteomes13030044","DOIUrl":"10.3390/proteomes13030044","url":null,"abstract":"<p><strong>Background: </strong>Mechanical damage to fruit during harvesting is nearly inevitable, with certain species, such as papaya, being particularly prone to spoilage. Postharvest handling can induce mechanical injuries that impair ripening and reduce shelf life, leading to significant economic losses. Although several studies have shed light on the molecular bases of mechanical damage, other aspects remain to be described (plant hormone inter-talk, physiological changes, and regulatory networks).</p><p><strong>Methods: </strong>In this study, we investigated proteomic changes in papaya fruit at two distinct ripening stages following mechanical damage. A total of 3230 proteins were identified, representing the most comprehensive proteomic analysis of papaya to date and the first assessment of proteins regulated by mechanical stress.</p><p><strong>Results: </strong>Proteins involved in ethylene biosynthesis were up-regulated on Day 2 but down-regulated on Day 12, with a similar trend observed for proteins in the abscisic acid synthesis pathway. Enzymes associated with photosynthesis, carbon fixation, primary metabolism, and carotenoid synthesis were down-regulated at both stages. In contrast, those related to plasmodesmata, calcium signaling, kinases, pathogenesis, cell wall remodeling, and proteases were up-regulated.</p><p><strong>Conclusions: </strong>These findings are thoroughly discussed, and a general model of the events triggered by mechanical impact in papaya is proposed. Our results provide a comprehensive framework for understanding papaya's response to mechanical damage.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-09-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452570/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113735","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparative Analysis of Plasma Extracellular Vesicle Isolation Methods for Purity Assessment and Biomarker Discovery.","authors":"Alexandra T Star, Melissa Hewitt, Amanpreet Badhwar, Wen Ding, Tammy-Lynn Tremblay, Jennifer J Hill, William G Willmore, Jagdeep K Sandhu, Arsalan S Haqqani","doi":"10.3390/proteomes13030045","DOIUrl":"10.3390/proteomes13030045","url":null,"abstract":"<p><strong>Background: </strong>Extracellular vesicles (EVs) are an important source of blood biomarkers and are emerging as next-generation therapeutics. Demonstrating the purity of isolated EVs is essential for applications ranging from proteomics-based biomarker discovery to biomanufacturing. In this study, we systematically evaluated multiple EV isolation methods for plasma and developed a scoring method to identify the approach best suited for proteomics.</p><p><strong>Methods: </strong>Commonly used enrichment techniques, including size-exclusion chromatography (SEC) and precipitation-based methods, were compared against the starting plasma in terms of particle yield and size, proteomic overlap, depletion of abundant plasma proteins, and enrichment of EV markers and unique proteins. To enable rigorous purity assessment, we established a targeted parallel reaction monitoring (PRM) mass spectrometry assay that quantified key EV markers and contaminant proteins across preparations.</p><p><strong>Results: </strong>Among the methods tested, SEC showed the greatest enrichment of EV markers and unique proteins, with the lowest level of contaminants, resulting in the highest overall purity scores. SEC also allowed for the detection of EV-free proteins. Other methods, by contrast, performed sub-optimally and were less reliable for proteomics-driven biomarker discovery.</p><p><strong>Conclusions: </strong>SEC provides the most EV-enriched plasma isolates for proteomics information, with minimal contamination from plasma proteins. The PRM-based purity scoring offers an objective means of benchmarking EV preparations and may help standardize EV isolation quality for both biomarker discovery and therapeutic manufacturing.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-09-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452325/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113778","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proteomic Analysis of Sputum from Patients with Active Tuberculosis.","authors":"Endrei Marcantonio, Amy M Woron, A Christian Whelen, Sladjana Prisic","doi":"10.3390/proteomes13030043","DOIUrl":"10.3390/proteomes13030043","url":null,"abstract":"<p><strong>Background: </strong>Patients with pulmonary tuberculosis (TB) typically produce sputa, which are used to identify the pathogen. Sputum also contains host proteins that may aid in diagnosis. We hypothesized that sputa from TB patients will have unique proteomes when compared to other lung diseases.</p><p><strong>Methods: </strong>Sputa were collected from 219 patients with suspected TB. Neutrophil-derived protein calprotectin (CP), which was used as a marker for lung damage, was quantified and compared between TB and non-TB groups. Three sputa with high or low CP from each group were selected and analyzed using label-free proteomics.</p><p><strong>Results: </strong>There was no difference in CP amounts between TB and non-TB groups. However, TB samples had other differentially abundant neutrophil-associated proteins. Compared to low CP, samples with high CP had much smaller number of proteins that could differentiate between TB and non-TB groups. Only two proteins, MUC5AC and MMP8, were more abundant in TB samples, regardless of CP levels.</p><p><strong>Conclusions: </strong>Our findings suggest that TB sputa may have unique proteomes that depend on CP levels, which should be further validated due to the small sample size. Therefore, controlled and more advanced TB may need a different set of biomarkers to reliably distinguish TB from other lung diseases.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452556/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113750","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proteomic Analysis of the Periodontal Ligament During Orthodontic Movement: A Study in Rats.","authors":"Camila Chierici Marcantonio, Maria Eduarda Scordamaia Lopes, Lélio Fernando Ferreira Soares, Cristiane Ribeiro Salmon, Francisco Humberto Nociti Junior, James Deschner, Andressa Vilas Boas Nogueira, Joni Augusto Cirelli","doi":"10.3390/proteomes13030042","DOIUrl":"10.3390/proteomes13030042","url":null,"abstract":"<p><p>The periodontal ligament (PDL) is a dynamic connective tissue that absorbs and transmits mechanical forces, playing a critical role during orthodontic tooth movement (OTM). This study aimed to characterize the proteomic profile of rat PDLs subjected to OTM. Ten Holtzman rats were allocated into Control and OTM groups. After 15 days of force application, hemimaxillae were harvested, and PDL tissues from the first maxillary molars were isolated via laser capture microdissection. Protein extracts were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS), followed by quantitative and enrichment analyses. Immunohistochemistry was performed to validate selected proteins. The full proteomic datasets supporting these findings are available in the PRIDE repository under the identifiers PXD055817 and PXD033647. A total of 1121 proteins were identified; 101 were exclusive to the OTM group, 324 to the control, and 696 shared. Among the 335 proteins with differential abundance, 334 were downregulated and one (Prelp) was upregulated in the OTM group. Enrichment analysis revealed that differentially abundant proteins were associated with molecular functions such as protein binding, and cellular components including extracellular exosomes, focal adhesions, and the extracellular matrix. Immunohistochemical analysis confirmed the presence of Prelp, Rbm3, and Cirbp in PDL tissues. These findings demonstrate that OTM significantly alters the proteomic landscape of the PDL and identify key proteins potentially involved in periodontal remodeling.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452640/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113794","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bidirectional Interaction Between PGE₂-Preconditioned Mesenchymal Stem Cells and Myofibroblasts Mediates Anti-Fibrotic Effects: A Proteomic Investigation into Equine Endometrial Fibrosis Reversal.","authors":"Lidice Méndez-Pérez, Yat Sen Wong, Belén O Ibáñez, Ioanna Martinez-Hormaza, Lleretny Rodríguez-Álvarez, Fidel Ovidio Castro","doi":"10.3390/proteomes13030041","DOIUrl":"10.3390/proteomes13030041","url":null,"abstract":"<p><strong>Background: </strong>Endometrosis is a prevalent fibrotic condition in mares that impairs reproductive efficiency by inducing transdifferentiation of endometrial stromal cells into myofibroblasts, leading to excessive ECM deposition.</p><p><strong>Methods: </strong>To elucidate the molecular mechanisms underlying fibrosis resolution, this study employed comprehensive proteomic techniques, including LC-MS/MS and SILAC, to analyze the interaction between myofibroblasts and mesenchymal stem cells derived from the endometrium (ET-eMSCs) preconditioned with PGE₂. An in vitro co-culture system was used, with samples collected at baseline and after 48 h.</p><p><strong>Results: </strong>Proteomic analysis identified significant alterations in proteins associated with ECM remodeling, immune regulation, and cellular stress response. Notably, proteins involved in collagen degradation, antioxidant defense, and growth factor signaling pathways were differentially abundant. Network analyses demonstrated robust interactions among these proteins, suggesting coordinated modulatory effects. The data indicate that PGE₂-primed ET-eMSCs induce a shift in myofibroblast secretory profiles, promoting a reduction in ECM stiffness, tissue reorganization, and activation of resolution pathways. Data are available via ProteomeXchange with identifier PXD067551.</p><p><strong>Conclusions: </strong>These findings reinforce the therapeutic potential of mesenchymal stem cell-based interventions for fibrotic diseases of the endometrium, opening avenues for regenerative strategies to restore reproductive function in mares.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452512/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113818","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Biological Variation in Serum ACE and CPN/CPB2 Activity in Healthy Individuals as Measured by the Degradation of Dabsylated Bradykinin-Reference Data and the Importance of Pre-Analytical Standardization.","authors":"Malte Bayer, Michael Snyder, Simone König","doi":"10.3390/proteomes13030040","DOIUrl":"10.3390/proteomes13030040","url":null,"abstract":"<p><strong>Background: </strong>Bradykinin (BK) is an inflammatory mediator. The degradation of labeled synthetic BK in biofluids can be used to report on the activity of angiotensin-converting enzyme (ACE) and basic carboxypeptidases N and CBP2, for which the neuropeptide is a substrate. Clinical studies have shown significant changes in the serum activity of these enzymes in patients with inflammatory diseases.</p><p><strong>Methods: </strong>Here, we investigated variation in the cleavage of dabsylated synthetic BK (DBK) in serum and the formation of the major enzymatic fragments using a thin-layer chromatography-based neuropeptide reporter assay (NRA) in a large cohort of healthy volunteers from the international human Personal Omics Profiling consortium based at Stanford University.</p><p><strong>Results: </strong>Four major outcomes were reported. First, a set of NRA reference data for the healthy population was delivered, which is important for future investigations of patient sera. Second, it was shown that the measured serum degradation capacity for DBK was significantly higher in males than in females. There was no significant correlation of the NRA results with ethnicity, body mass index or overnight fasting. Third, a batch effect was noted among sampling sites (HUPO conferences). Thus, we used subcohorts rather than the entire collection for data mining. Fourth, as the low-cost and robust NRA is sensitive to enzyme activity, it provides such a necessary quick test to eliminate degraded and/or otherwise questionable samples.</p><p><strong>Conclusions: </strong>The results reiterate the critical importance of a high level of standardization in pre-analytical sample collection and processing-most notably, sample quality should be evaluated before conducting any large and expensive omics analyses.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-08-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452422/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113903","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Adiposome Proteomics Uncover Molecular Signatures of Cardiometabolic Risk in Obese Individuals.","authors":"Mohamed Saad Rakab, Monica C Asada, Imaduddin Mirza, Mohammed H Morsy, Amro Mostafa, Francesco M Bianco, Mohamed M Ali, Chandra Hassan, Mario A Masrur, Brian T Layden, Abeer M Mahmoud","doi":"10.3390/proteomes13030039","DOIUrl":"10.3390/proteomes13030039","url":null,"abstract":"<p><strong>Background: </strong>Adipose-derived extracellular vesicles (adiposomes) are emerging as key mediators of inter-organ communication, yet their molecular composition and role in obesity-related pathophysiology remain underexplored. This study integrates clinical phenotyping with proteomic analysis of visceral adipose-derived adiposomes to identify obesity-linked molecular disruptions.</p><p><strong>Methods: </strong>Seventy-five obese and forty-seven lean adults were extensively profiled for metabolic, inflammatory, hepatic, and vascular parameters. Adiposomes isolated from visceral fat underwent mass spectrometry-based proteomic analysis, followed by differential abundance, pathway enrichment, regulatory network modeling, and clinical association testing.</p><p><strong>Results: </strong>Obese individuals exhibited widespread cardiometabolic dysfunction. Proteomics revealed 64 adiposomal proteins with differential abundance. Upregulated proteins (e.g., CRP, C9, APOC1) correlated with visceral adiposity, systemic inflammation, and endothelial dysfunction. In contrast, downregulated proteins (e.g., ADIPOQ, APOD, TTR, FGB, FGG) were associated with enhanced nitric oxide bioavailability and vascular protection, suggesting loss of homeostatic signaling. Network analyses identified TNF and IL1 as key upstream regulators driving inflammatory and oxidative stress pathways. Decision tree and random forest models accurately classified obesity, hypertension, diabetes, dyslipidemia, and hepatic steatosis (AUC = 0.908-0.994), identifying predictive protein signatures related to complement activation, inflammation, and lipid transport.</p><p><strong>Conclusion: </strong>Obesity alters adiposome proteomic cargo, reflecting and potentially mediating systemic inflammation, metabolic dysregulation, and vascular impairment.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-08-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12452415/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145113831","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of Cancer-Associated Proteins in Colorectal Cancer Using Mass Spectrometry.","authors":"Naoyuki Toyota, Ryo Konno, Shuhei Iwata, Shin Fujita, Yoshio Kodera, Rei Noguchi, Tadashi Kondo, Yusuke Kawashima, Yuki Yoshimatsu","doi":"10.3390/proteomes13030038","DOIUrl":"10.3390/proteomes13030038","url":null,"abstract":"<p><strong>Background: </strong>Colorectal cancer (CRC) is a leading cause of cancer-related mortality worldwide, with a multifactorial etiology involving genetic and environmental factors. Advanced proteomics offers valuable insights into the molecular mechanisms of cancer, identifying proteins that function as mediators in tumor biology.</p><p><strong>Methods: </strong>In this study, we used mass spectrometry-based data-independent acquisition (DIA) to analyze the proteomic landscape of CRC. We compared protein abundance in normal and tumor tissues from 16 patients with CRC to identify cancer-associated proteins and examine their roles in disease progression.</p><p><strong>Results: </strong>The analysis identified 10,329 proteins, including 531 cancer-associated proteins from the Catalogue Of Somatic Mutations In Cancer (COSMIC) database, and 48 proteins specifically linked to CRC. Notably, clusters of proteins showed consistent increases or decreases in abundance across disease stages, suggesting their roles in tumorigenesis and progression.</p><p><strong>Conclusions: </strong>Our findings suggest that proteome abundance trends may contribute to the identification of biomarker candidates and therapeutic targets in colorectal cancer. However, given the limited sample size and lack of subtype stratification, further studies using larger, statistically powered cohorts are warranted to establish clinical relevance. These proteins may provide insights into drug resistance and tumor heterogeneity. Limitations of the study include the inability to detect low-abundance proteins and reliance on protein abundance rather than functional activity. Future complementary approaches, such as affinity proteomics, are suggested to address these limitations.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12372073/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144966633","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Uncovering Enzyme-Specific Post-Translational Modifications: An Overview of Current Methods.","authors":"Nashira H Ridgeway, Kyle K Biggar","doi":"10.3390/proteomes13030037","DOIUrl":"10.3390/proteomes13030037","url":null,"abstract":"<p><p>Post-translational modifications (PTMs) govern a multitude of protein functions within the cell, surpassing the basic function(s) encoded directly within the amino acid sequence. Despite the historical discovery of PTMs dating back over a century, recent technological advancements have facilitated the rapid expansion of the known PTM landscape. However, the elucidation of enzyme-substrate relationships responsible for PTMs, particularly for those less studied, remains a challenging endeavor. This review provides an extensive overview of methods employed in the discovery of enzyme-specific substrates for PTM catalysis. Beginning with traditional experimental approaches rooted in chemistry, biochemistry and cell biology, this review progresses to recently developed computational strategies tailored for identifying enzyme-substrate interactions. The analysis reflects on the remarkable progress achieved in PTM research to date, underscoring the increasing role of computational and high-throughput techniques in expediting enzyme-substrate discovery. Furthermore, it highlights the potential of artificial intelligence to revolutionize PTM research and emphasizes the importance of unbiased high-throughput analysis in advancing our understanding of PTM networks. Ultimately, the review advocates for the integration of sophisticated computational strategies with experimental techniques to unravel the complex enzyme-substrate networks governing PTM-mediated cellular processes.</p>","PeriodicalId":20877,"journal":{"name":"Proteomes","volume":"13 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-08-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12371983/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144966574","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}