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Fluorescence Localization of the Conjugation Machinery of Bacillus subtilis
Author(s) -
Chen Sirui,
Dame Haley B.,
Berkmen Melanie B.
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.00626
Subject(s) - green fluorescent protein , mcherry , bacillus subtilis , subcellular localization , fusion protein , microbiology and biotechnology , transmembrane protein , protein subcellular localization prediction , biology , inner membrane , transport protein , secretion , biophysics , membrane , biochemistry , gene , bacteria , genetics , recombinant dna , receptor , cytoplasm
Conjugation, or mating, is the transfer of DNA from a donor to a recipient cell through conjugation machinery. Conjugation contributes to bacterial genome innovation and the spread of virulence and antibiotic resistance genes between bacteria. DNA is transferred during conjugation through a multi‐protein DNA translocation channel, classified as a Type IV secretion system (T4SS), embedded in the membrane. Our work focuses on characterizing the conjugation machinery of the integrative and conjugative element ICE Bs1 of Bacillus subtilis . Previously, we identified several protein components of the ICE Bs1 conjugation machinery, including the bitopic membrane protein ConB and the peripheral membrane ATPase ConE. We have used fluorescence microscopy to observe the locations of ConB and ConE inside cells. A fusion of ConE to green fluorescent protein (GFP) localizes to the membrane, predominantly at the cell poles. Here, we show that a fusion of ConB to the red fluorescent protein mCHERRY localizes similarly to ConE‐GFP. Consistent with ConE lacking predicted transmembrane segments, ConE‐GFP’s localization to the membrane requires ConB. In contrast, localization of ConB‐mCHERRY to the membrane does not depend on other conjugation proteins. Current studies are focused on which domains of ConB are required for its own localization as well as the localization of ConE. Support or Funding Information This research was funded by Suffolk University and an NSF‐RUI grant to M. Berkmen.