Genetic Dissection of a Prevalent Plasmid-Encoded Conjugation System in

Overview

Journal Front Microbiol

Specialty Microbiology

Date 2021 Jun 14

PMID 34122391

Citations 10

Authors

Guillermo Ortiz Charneco

Philip Kelleher

Andrius Buivydas

Hugo Streekstra

Emiel Ver Loren van Themaat

Paul P de Waal

Jennifer Mahony

Douwe Van Sinderen

Affiliations

Soon will be listed here.

Abstract

Plasmid pNP40, which was first identified nearly 40 years ago in subsp. biovar diacetylactis DRC3, encodes functions such as heavy metal-, bacteriophage-, and nisin-resistance, as well as plasmid transfer ability by conjugation. Here, we report an optimized conjugation protocol for this plasmid, yielding a transfer frequency that is approximately 4,000-fold higher than those previously reported in literature, while we also observed high-frequency plasmid co-mobilization. Individual mutations in 18 genes that encompass the presumed conjugation cluster of pNP40 were generated using ssDNA recombineering to evaluate the role of each gene in the conjugation process. A possible transcriptional repressor of this conjugation cluster, the product of the R gene, was identified in this manner. This mutational analysis, paired with bioinformatic predictions as based on sequence and structural similarities, allowed us to generate a preliminary model of the pNP40 conjugation machinery.

Citing Articles

Functional and practical insights into three lactococcal antiphage systems.

Grafakou A, Mosterd C, de Waal P, van Rijswijck I, van Peij N, Mahony J Appl Environ Microbiol. 2024; 90(9):e0112024.

PMID: 39136492 PMC: 11409693. DOI: 10.1128/aem.01120-24.

Discovery of antiphage systems in the lactococcal plasmidome.

Grafakou A, Mosterd C, Beck M, Kelleher P, McDonnell B, de Waal P Nucleic Acids Res. 2024; 52(16):9760-9776.

PMID: 39119896 PMC: 11381338. DOI: 10.1093/nar/gkae671.

Plasmid-mediated horizontal gene mobilisation: Insights from two lactococcal conjugative plasmids.

Ortiz Charneco G, McDonnell B, Kelleher P, Buivydas A, Dashko S, de Waal P Microb Biotechnol. 2024; 17(5):e14421.

PMID: 38752994 PMC: 11097999. DOI: 10.1111/1751-7915.14421.

Discovering genetic determinants for cell-to-cell adhesion in two prevalent conjugative lactococcal plasmids.

Ortiz Charneco G, Kelleher P, Buivydas A, de Waal P, van Rijswijck I, van Peij N Curr Res Microb Sci. 2024; 6:100239.

PMID: 38706493 PMC: 11067333. DOI: 10.1016/j.crmicr.2024.100239.

Effect of TraN key residues involved in DNA binding on pIP501 transfer rates in .

Michaelis C, Berger T, Kuhlmann K, Ghulam R, Petrowitsch L, Besora Vecino M Front Mol Biosci. 2024; 11:1268647.

PMID: 38380428 PMC: 10877727. DOI: 10.3389/fmolb.2024.1268647.

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