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oai:figshare.com:article/34025004 2026-09-29T16:06:16Z category_1 category_4 category_8 category_13 category_16 category_272 category_873 category_734 portal_63 item_type_6 month_year_09_2026

Proteomics-Guided Discovery of Candidate Enzymes for Degradation of a Rare Pyruvylated Glycosaminogalactan by a Newly Isolated Luteolibacter Strain Victor Daisuke Kietzmann (25137807) Pascal M. Mrozek (25137810) G. Jacob Boehlich (25137813) Ronja M. Sandholm (25137816) Sabina L. La Rosa (25137819) Bjørge Westereng (338804) Gustav Vaaje-Kolstad (188321) Biophysics Biochemistry Microbiology Genetics Physiology Environmental Sciences not elsewhere classified Chemical Sciences not elsewhere classified Biological Sciences not elsewhere classified strain bacterial exopolysaccharides related structural homologues pyruvylated bacterial glycan novel strain capable distantly related gh123 based structural annotation await experimental validation acting cleavage followed secreted gh123 protein rare pyruvylated glycosaminogalactan ></ sub >, luteolibacter </ italic vk1 depolymerizes eps mass glycosaminogalactan luteolibacter </ like protein ></ sub sub >< structurally complex strongly associated r </ newly isolated monosaccharide composition may contribute lp </ limited coverage guided discovery glycosidic linkages gal motif findings support exclusion chromatography enzymes responsible enzyme family enrichment cultivation defined oligosaccharides current carbohydrate combining genomics Bacterial exopolysaccharides (EPS) display extensive structural diversity arising from variation in monosaccharide composition, glycosidic linkages, and non-carbohydrate modifications. However, the enzymes responsible for degrading many EPS structures remain unknown, reflecting the limited coverage of current carbohydrate-active enzyme (CAZyme) families. Here, we investigated the microbial degradation of EPS_<i>Lp</i>, a high-molecular-mass glycosaminogalactan from Lactiplantibacillus pentosus KW1 containing the uncommon GalNAcβ1 → 2(4,6-<i>O</i>-<i>R</i>-pyruvyl)Gal motif. Enrichment cultivation with EPS_<i>Lp</i> as the main carbon source led to the isolation of <i>Luteolibacter</i> sp. VK1, a novel strain capable of utilizing this biopolymer. Size-exclusion chromatography and MALDI-TOF mass spectrometry showed that VK1 depolymerizes EPS_<i>Lp</i> into defined oligosaccharides, consistent with endo-acting cleavage followed by further processing. By combining genomics, AlphaFold-based structural annotation and comparative proteomics, we identified candidate CAZymes potentially involved in EPS_<i>Lp</i> catabolism. Notably, a secreted GH123 protein and a distantly related GH123-like protein were strongly associated with EPS_<i>Lp</i>-dependent growth, suggesting that this enzyme family, or related structural homologues, may contribute to extracellular EPS_<i>Lp</i> depolymerization. Together, these findings support a candidate pathway for degradation of a structurally complex, pyruvylated bacterial glycan and highlight potentially novel EPS-active enzymes whose proposed roles, predicted through bioinformatic analyses, await experimental validation. 2026-09-29T00:00:00Z Text Journal contribution 10.1021/acs.jproteome.6c00586.s001 https://figshare.com/articles/journal_contribution/Proteomics-Guided_Discovery_of_Candidate_Enzymes_for_Degradation_of_a_Rare_Pyruvylated_Glycosaminogalactan_by_a_Newly_Isolated_Luteolibacter_Strain/34025004 CC BY-NC 4.0