{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T18:38:52Z","timestamp":1790620732235,"version":"4.1.0"},"posted":{"date-parts":[[2024,8,7]]},"group-title":"Microbiology","reference-count":66,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2024,8,7]],"date-time":"2024-08-07T00:00:00Z","timestamp":1722988800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,8,6]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>\n                  In diderm bacteria, the Lol pathway canonically mediates the periplasmic transport of lipoproteins from the inner membrane (IM) to the outer membrane (OM) and therefore plays an essential role in bacterial envelope homeostasis. After extrusion of modified lipoproteins from the IM via the LolCDE complex, the periplasmic chaperone LolA carries lipoproteins through the periplasm and transfers them to the OM lipoprotein insertase LolB, itself a lipoprotein with a LolA-like fold. Yet, LolB homologs appear restricted to \u03c8-proteobacteria and are missing from spirochetes like the tick-borne Lyme disease pathogen\n                  <jats:italic>Borrelia burgdorferi<\/jats:italic>\n                  , suggesting a different hand-off mechanism at the OM. Here, we solved the crystal structure of the\n                  <jats:italic>B. burgdorferi<\/jats:italic>\n                  LolA homolog BB0346 (LolA\n                  <jats:sub>Bb<\/jats:sub>\n                  ) at 1.9 \u00c5 resolution. We identified multiple structural deviations in comparative analyses to other solved LolA structures, particularly a unique LolB-like protruding loop domain. LolA\n                  <jats:sub>Bb<\/jats:sub>\n                  failed to complement an\n                  <jats:italic>Escherichia coli lolA<\/jats:italic>\n                  knockout, even after codon optimization, signal I peptide adaptation, and a C-terminal chimerization which had allowed for complementation with an \u03b1-proteobacterial LolA. Analysis of a conditional\n                  <jats:italic>B. burgdorferi lolA<\/jats:italic>\n                  knockout strain indicated that LolA\n                  <jats:sub>Bb<\/jats:sub>\n                  was essential for growth. Intriguingly, protein localization assays indicated that initial depletion of LolA\n                  <jats:sub>Bb<\/jats:sub>\n                  led to an emerging mislocalization of both IM and periplasmic OM lipoproteins, but not surface lipoproteins. Together, these findings further support the presence of two separate primary secretion pathways for periplasmic and surface OM lipoproteins in\n                  <jats:italic>B. burgdorferi<\/jats:italic>\n                  and suggest that the distinct structural features of LolA\n                  <jats:sub>Bb<\/jats:sub>\n                  allow it to function in a unique LolB-deficient lipoprotein sorting system.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>SIGNIFICANCE<\/jats:title>\n                  <jats:p>\n                    <jats:italic>Borrelia<\/jats:italic>\n                    spirochetes causing Lyme disease and relapsing fever have unusual double-membrane envelopes that instead of lipopolysaccharide (LPS) display abundant surface lipoproteins. We recently showed that secretion of these surface lipoproteins in\n                    <jats:italic>Borrelia burgdorferi<\/jats:italic>\n                    depends on a distant homolog of the canonical LPS outer membrane translocase LptD. Here, we probed the role of the\n                    <jats:italic>B. burgdorferi<\/jats:italic>\n                    Lol pathway in lipoprotein sorting and secretion. We show that the periplasmic chaperone LolA is essential, functionally different from\n                    <jats:italic>E. coli<\/jats:italic>\n                    LolA, with structural features of a bifunctional lipoprotein carrier protein operating without a downstream LolB outer membrane lipoprotein insertase. Depletion of LolA did not impact surface lipoprotein localization but led to a marked mislocalization of inner membrane lipoproteins to the outer membrane. This further supports two parallel, yet potentially interacting\n                    <jats:italic>Borrelia<\/jats:italic>\n                    lipoprotein transport pathways that are responsible for either secreting surface lipoprotein virulence factors or maintaining proper distribution of lipoproteins within the periplasmic space.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.1101\/2024.08.06.606844","type":"posted-content","created":{"date-parts":[[2024,8,7]],"date-time":"2024-08-07T10:15:16Z","timestamp":1723025716000},"source":"Crossref","is-referenced-by-count":4,"title":["<i>Borrelia burgdorferi<\/i>\n                  BB0346 is an Essential, Structurally Variant LolA Homolog that is Primarily Required for Homeostatic Localization of Periplasmic Lipoproteins"],"prefix":"10.64898","author":[{"given":"Bryan T.","family":"Murphy","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jacob J.","family":"Wiepen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Danielle E.","family":"Graham","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Selene K.","family":"Swanson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maithri M.","family":"Kashipathy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anne","family":"Cooper","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kevin P.","family":"Battaile","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David K.","family":"Johnson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Laurence","family":"Florens","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6216-7365","authenticated-orcid":false,"given":"Jon S.","family":"Blevins","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Scott","family":"Lovell","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7350-2162","authenticated-orcid":false,"given":"Wolfram R.","family":"Z\u00fcckert","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2024080812300960000_2024.08.06.606844v1.1","doi-asserted-by":"crossref","unstructured":"Z\u00fcckert WR . 2019. 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