{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T11:34:57Z","timestamp":1785411297176,"version":"3.56.0"},"posted":{"date-parts":[[2025,6,25]]},"group-title":"Microbiology","reference-count":84,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,6,25]],"date-time":"2025-06-25T00:00:00Z","timestamp":1750809600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"award":["R01-AI155915"],"award-info":[{"award-number":["R01-AI155915"]}],"id":[{"id":"https:\/\/ror.org\/043z4tv69","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,12,10]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Enterobacterial common antigen (ECA) is a conserved glycan that supports outer membrane impermeability and intrinsic antibiotic resistance in Enterobacterales. ECA exists in outer membrane diacylglycerol-phosphate- and lipopolysaccharide-linked forms, and a cyclic periplasmic form (ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  ). Intriguingly, ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  both affects the outer-membrane permeability barrier and functions in regulation of diacylglycerol-phosphate-linked ECA levels. While the length of linear ECA polymers generated by WzyE is regulated by the co-polymerase WzzE, WzzE is also required for ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  biogenesis and no ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  is synthesized in its absence. To uncover WzzE functions necessary for ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  biosynthesis, we generated plasmid-borne and chromosomal\n                  <jats:italic>wzzE<\/jats:italic>\n                  mutants in\n                  <jats:italic>Escherichia coli<\/jats:italic>\n                  K-12 and quantified their effects on linear chain-length regulation and on ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  synthesis. Mutations affecting linear ECA chain-length regulation in either transmembrane helix 2 or the periplasmic domain abolished ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  synthesis. Notably, two variants at residue F104 exhibited identical linear ECA chain-length regulation which was similar to wild type, but resulted in sharply different ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  production: chromosomal WzzE\n                  <jats:sub>F104Y<\/jats:sub>\n                  produced near wild-type levels of ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  , whereas WzzE\n                  <jats:sub>F104H<\/jats:sub>\n                  produced approximately two-fold less ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  . This difference in ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  production vs. linear chain-length regulation demonstrates ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  synthesis is genetically separable from this regulation. Thus, WzzE-mediated chain-length regulation is necessary for ECA\n                  <jats:sub>CYC<\/jats:sub>\n                  biogenesis, but not sufficient.\n                <\/jats:p>","DOI":"10.1101\/2025.06.25.661564","type":"posted-content","created":{"date-parts":[[2025,6,26]],"date-time":"2025-06-26T01:00:10Z","timestamp":1750899610000},"source":"Crossref","is-referenced-by-count":1,"title":["Chain-length regulation by WzzE is necessary for, but genetically separable from, cyclic enterobacterial common antigen synthesis"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9228-8168","authenticated-orcid":false,"given":"Joseph F.","family":"Carr","sequence":"first","affiliation":[{"name":"Department of Biology, Texas A&M University, College Station, Texas, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jennifer S.","family":"Rudolf","sequence":"additional","affiliation":[{"name":"Department of Biology, Texas A&M University, College Station, Texas, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Daniel J.","family":"Warzecha","sequence":"additional","affiliation":[{"name":"Department of Biology, Texas A&M University, College Station, Texas, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yohannes H.","family":"Rezenom","sequence":"additional","affiliation":[{"name":"Chemistry Mass Spectrometry Facility, Texas A&M University, College Station, Texas, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1062-0716","authenticated-orcid":false,"given":"Angela M.","family":"Mitchell","sequence":"additional","affiliation":[{"name":"Department of Biology, Texas A&M University, College Station, Texas, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026041210100674000_2025.06.25.661564v2.1","doi-asserted-by":"publisher","DOI":"10.1038\/S41586-024-07487-W"},{"key":"2026041210100674000_2025.06.25.661564v2.2","doi-asserted-by":"publisher","DOI":"10.1099\/ijsem.0.001485"},{"key":"2026041210100674000_2025.06.25.661564v2.3","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.00315-16"},{"key":"2026041210100674000_2025.06.25.661564v2.4","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2010.00134"},{"key":"2026041210100674000_2025.06.25.661564v2.5","doi-asserted-by":"publisher","DOI":"10.1107\/S090744499901481X"},{"key":"2026041210100674000_2025.06.25.661564v2.6","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1006434"},{"key":"2026041210100674000_2025.06.25.661564v2.7","first-page":"e00224","article-title":"Identification of the Shigella flexneri Wzy Domain Modulating Wzz(pHS-2) Interaction and Detection of the Wzy\/Wzz\/Oag Complex","volume":"204","year":"2022","journal-title":"J Bacteriol"},{"key":"2026041210100674000_2025.06.25.661564v2.8","doi-asserted-by":"publisher","DOI":"10.1038\/msb4100050"},{"key":"2026041210100674000_2025.06.25.661564v2.9","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(18)80083-7"},{"key":"2026041210100674000_2025.06.25.661564v2.10","doi-asserted-by":"publisher","DOI":"10.1128\/JB.181.20.6564-6568.1999"},{"key":"2026041210100674000_2025.06.25.661564v2.11","doi-asserted-by":"publisher","DOI":"10.1128\/jb.171.3.1326-1332.1989"},{"key":"2026041210100674000_2025.06.25.661564v2.12","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(18)48216-6"},{"key":"2026041210100674000_2025.06.25.661564v2.13","doi-asserted-by":"publisher","DOI":"10.1111\/j.1432-1033.1987.tb10544.x"},{"key":"2026041210100674000_2025.06.25.661564v2.14","doi-asserted-by":"crossref","first-page":"250055","DOI":"10.1098\/rsob.250055","article-title":"Recent advances in understanding of enterobacterial common antigen synthesis and regulation","volume":"15","year":"2025","journal-title":"Open Biology"},{"key":"2026041210100674000_2025.06.25.661564v2.15","first-page":"ESP-0001-2018","article-title":"Function and Biogenesis of Lipopolysaccharides","volume":"8","year":"2018","journal-title":"EcoSal Plus"},{"key":"2026041210100674000_2025.06.25.661564v2.16","doi-asserted-by":"publisher","DOI":"10.1016\/j.str.2017.03.017"},{"key":"2026041210100674000_2025.06.25.661564v2.17","doi-asserted-by":"publisher","DOI":"10.1046\/j.1365-2958.1999.01591.x"},{"key":"2026041210100674000_2025.06.25.661564v2.18","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.120163297"},{"key":"2026041210100674000_2025.06.25.661564v2.19","unstructured":"Doern, C. 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