In:
PLOS ONE, Public Library of Science (PLoS), Vol. 15, No. 11 ( 2020-11-23), p. e0242593-
Abstract:
Bacterial efflux pumps are an important pathogenicity trait because they extrude a variety of xenobiotics. Our laboratory previously identified in silico Burkholderia collagen-like protein 8 (Bucl8) in the hazardous pathogens Burkholderia pseudomallei and Burkholderia mallei . We hypothesize that Bucl8, which contains two predicted tandem outer membrane efflux pump domains, is a component of a putative efflux pump. Unique to Bucl8, as compared to other outer membrane proteins, is the presence of an extended extracellular region containing a collagen-like (CL) domain and a non-collagenous C-terminus (Ct). Molecular modeling and circular dichroism spectroscopy with a recombinant protein, corresponding to this extracellular CL-Ct portion of Bucl8, demonstrated that it adopts a collagen triple helix, whereas functional assays screening for Bucl8 ligands identified binding to fibrinogen. Bioinformatic analysis of the bucl8 gene locus revealed it resembles a classical efflux-pump operon. The bucl8 gene is co-localized with downstream fusCDE genes encoding fusaric acid (FA) resistance, and with an upstream gene, designated as fusR , encoding a LysR-type transcriptional regulator. Using reverse transcriptase (RT)-qPCR, we defined the boundaries and transcriptional organization of the fusR-bucl8-fusCDE operon. We found exogenous FA induced bucl8 transcription over 80-fold in B . pseudomallei , while deletion of the entire bucl8 locus decreased the minimum inhibitory concentration of FA 4-fold in its isogenic mutant. We furthermore showed that the putative Bucl8-associated pump expressed in the heterologous Escherichia coli host confers FA resistance. On the contrary, the Bucl8-associated pump did not confer resistance to a panel of clinically-relevant antimicrobials in Burkholderia and E . coli . We finally demonstrated that deletion of the bucl8 -locus drastically affects the growth of the mutant in L-broth. We determined that Bucl8 is a component of a novel tetrapartite efflux pump, which confers FA resistance, fibrinogen binding, and optimal growth.
Type of Medium:
Online Resource
ISSN:
1932-6203
DOI:
10.1371/journal.pone.0242593
DOI:
10.1371/journal.pone.0242593.g001
DOI:
10.1371/journal.pone.0242593.g002
DOI:
10.1371/journal.pone.0242593.g003
DOI:
10.1371/journal.pone.0242593.g004
DOI:
10.1371/journal.pone.0242593.g005
DOI:
10.1371/journal.pone.0242593.g006
DOI:
10.1371/journal.pone.0242593.g007
DOI:
10.1371/journal.pone.0242593.t001
DOI:
10.1371/journal.pone.0242593.t002
DOI:
10.1371/journal.pone.0242593.t003
DOI:
10.1371/journal.pone.0242593.s001
DOI:
10.1371/journal.pone.0242593.s002
DOI:
10.1371/journal.pone.0242593.s003
DOI:
10.1371/journal.pone.0242593.s004
DOI:
10.1371/journal.pone.0242593.s005
Language:
English
Publisher:
Public Library of Science (PLoS)
Publication Date:
2020
detail.hit.zdb_id:
2267670-3
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