The Interaction of the F-Like Plasmid-Encoded TraN Isoforms with Their Cognate Outer Membrane Receptors

Abstract

Conjugation plays a central role in the spread of antimicrobial resistance genes among bacterial pathogens. Efficient conjugation is mediated by formation of mating pairs via a pilus, followed by mating pair stabilization (MPS), mediated by tight interactions between the plasmid-encoded outer membrane protein (OMP) TraN in the donor (of which there are 7 sequence types grouped into the 4 structural isoforms α, β, γ, and δ), and an OMP receptor in the recipient (OmpW, OmpK36, OmpA, and OmpF, respectively)

A Whole-Cell Screening Platform to Discover Cell Adhesion Molecules that Enable Programmable Bacterial Cell-Cell Adhesion

“…The bacteria nanobody display library (100 ng) was then transformed into the E. coli MG1655 ΔompW strain via electroporation. We selected the Δ ompW strain to minimize the interactions between OmpW and TraN, as suggested by Low et al 21, 41 . The cells were harvested and resuspended in LB with 10% glycerol for storage at -80 °C.…”

Structure of the outer membrane porin OmpW from the pervasive pathogen Klebsiella pneumoniae

“…Despite amino-acid differences in the extracellular region between OmpW KP and OmpW EC (Fig. 3c), where the tip of TraN R100-1 has been shown to bind (Low et al, 2023), binding of TraN R100-1 is not impaired between the two species. We previously reported that Ala142, which is conserved between OmpW KP and OmpW EC , acts as the minimum residue for specificity towards TraN R100-1 (Low et al, 2023); the equivalent residue in Citrobacter rodentium OmpW (OmpW CR ) is Asn142, which prevents R100-1 conjugation because of a steric clash with the tip of TraN R100-1 (Low et al, 2023).…”

“…We previously reported that Ala142, which is conserved between OmpW KP and OmpW EC , acts as the minimum residue for specificity towards TraN R100-1 (Low et al, 2023); the equivalent residue in Citrobacter rodentium OmpW (OmpW CR ) is Asn142, which prevents R100-1 conjugation because of a steric clash with the tip of TraN R100-1 (Low et al, 2023). The N142A mutation in OmpW CR restored conjugation efficiency (Low et al, 2023).…”

“…It has also been shown that OMPs participate in F-like plasmid conjugation, a form of horizontal gene transfer where plasmids are transferred from donor to recipient bacteria in a contact-dependent manner (Lederberg & Tatum, 1946;Frankel et al, 2023). We have recently shown that the efficient conjugation of the multidrugresistant R100-1 plasmid into both Escherichia coli (EC) and Klebsiella pneumoniae (KP) relies on the formation of matingpair stabilization via interaction between the R100-1-encoded OM protein TraN� in the donor and the OMP OmpW EC or OmpW KP in the recipient (Low et al, 2022(Low et al, , 2023. Pairing of the TraN isoform with recipient receptors mediates conjugation species specificity and host range; an in-depth review of mating-pair stabilization and the role of TraN has been provided by Frankel et al (2023).…”

De Novo Synthesis of a Conjugative System from Human Gut Metagenomic Data for Targeted Delivery of Cas9 Antimicrobials

“…Conjugative plasmids are well suited as delivery vectors for CRISPR nucleases or other genetic tools; they have a large coding capacity, they often encode CRISPR systems, , they encode factors that promote biofilm formation to enhance cell-to-cell contact and DNA transfer, they can carry antirestriction systems to promote resistance to restriction-modification systems, − and they generally do not require a cellular receptor that would provide a mechanism for bacterial resistance, although some conjugative systems do require a receptor. , Conjugative systems initiate the unidirectional transfer of DNA from a donor to a recipient cell through the actions of the relaxase protein (MOB) that nicks DNA at a defined origin of the transfer sequence ( oriT ). , Subsequent interaction of the relaxosome with the type IV coupling protein (T4CP), mating pair formation, and type 4 secretion (T4SS) proteins catalyze DNA transfer to the recipient. − Conjugative systems can exist as complete systems that are self-transmissible or as partial systems that are dependent on conjugative proteins encoded by other elements. Upon DNA transfer, some conjugative systems integrate into the chromosome as integrative conjugative elements (ICEs). − Currently, the classification of conjugative systems is based on the identity of the MOB protein …”