Crystal structure of TcpK in complex with oriT DNA of the antibiotic resistance plasmid pCW3

Daouda A.K. Traore, Jessica A. Wisniewski, Sarena F. Flanigan, Paul J. Conroy, Santosh Panjikar, Yee-Foong Mok, Carmen Lao, Michael D.W. Griffin, Vicki Adams, Julian I. Rood, James C. Whisstock

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Conjugation is fundamental for the acquisition of new genetic traits and the development of antibiotic resistance in pathogenic organisms. Here, we show that a hypothetical Clostridium perfringens protein, TcpK, which is encoded by the tetracycline resistance plasmid pCW3, is essential for efficient conjugative DNA transfer. Our studies reveal that TcpK is a member of the winged helix-turn-helix (wHTH) transcription factor superfamily and that it forms a dimer in solution. Furthermore, TcpK specifically binds to a nine-nucleotide sequence that is present as tandem repeats within the pCW3 origin of transfer (oriT). The X-ray crystal structure of the TcpK–TcpK box complex reveals a binding mode centered on and around the β-wing, which is different from what has been previously shown for other wHTH proteins. Structure-guided mutagenesis experiments validate the specific interaction between TcpK and the DNA molecule. Additional studies highlight that the TcpK dimer is important for specific DNA binding.

Original languageEnglish
Article number3732
Number of pages11
JournalNature Communications
Volume9
Issue number1
DOIs
Publication statusPublished - 13 Sep 2018

Keywords

  • bacterial genetics
  • DNA
  • x-ray crystallography

Cite this

@article{2c6ccfacd05b41acab20fe4aaf5d4215,
title = "Crystal structure of TcpK in complex with oriT DNA of the antibiotic resistance plasmid pCW3",
abstract = "Conjugation is fundamental for the acquisition of new genetic traits and the development of antibiotic resistance in pathogenic organisms. Here, we show that a hypothetical Clostridium perfringens protein, TcpK, which is encoded by the tetracycline resistance plasmid pCW3, is essential for efficient conjugative DNA transfer. Our studies reveal that TcpK is a member of the winged helix-turn-helix (wHTH) transcription factor superfamily and that it forms a dimer in solution. Furthermore, TcpK specifically binds to a nine-nucleotide sequence that is present as tandem repeats within the pCW3 origin of transfer (oriT). The X-ray crystal structure of the TcpK–TcpK box complex reveals a binding mode centered on and around the β-wing, which is different from what has been previously shown for other wHTH proteins. Structure-guided mutagenesis experiments validate the specific interaction between TcpK and the DNA molecule. Additional studies highlight that the TcpK dimer is important for specific DNA binding.",
keywords = "bacterial genetics, DNA, x-ray crystallography",
author = "Traore, {Daouda A.K.} and Wisniewski, {Jessica A.} and Flanigan, {Sarena F.} and Conroy, {Paul J.} and Santosh Panjikar and Yee-Foong Mok and Carmen Lao and Griffin, {Michael D.W.} and Vicki Adams and Rood, {Julian I.} and Whisstock, {James C.}",
year = "2018",
month = "9",
day = "13",
doi = "10.1038/s41467-018-06096-2",
language = "English",
volume = "9",
journal = "Nature Communications",
issn = "2041-1723",
publisher = "Nature Publishing Group",
number = "1",

}

Crystal structure of TcpK in complex with oriT DNA of the antibiotic resistance plasmid pCW3. / Traore, Daouda A.K.; Wisniewski, Jessica A.; Flanigan, Sarena F.; Conroy, Paul J.; Panjikar, Santosh; Mok, Yee-Foong; Lao, Carmen; Griffin, Michael D.W.; Adams, Vicki; Rood, Julian I.; Whisstock, James C.

In: Nature Communications, Vol. 9, No. 1, 3732, 13.09.2018.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Crystal structure of TcpK in complex with oriT DNA of the antibiotic resistance plasmid pCW3

AU - Traore, Daouda A.K.

AU - Wisniewski, Jessica A.

AU - Flanigan, Sarena F.

AU - Conroy, Paul J.

AU - Panjikar, Santosh

AU - Mok, Yee-Foong

AU - Lao, Carmen

AU - Griffin, Michael D.W.

AU - Adams, Vicki

AU - Rood, Julian I.

AU - Whisstock, James C.

PY - 2018/9/13

Y1 - 2018/9/13

N2 - Conjugation is fundamental for the acquisition of new genetic traits and the development of antibiotic resistance in pathogenic organisms. Here, we show that a hypothetical Clostridium perfringens protein, TcpK, which is encoded by the tetracycline resistance plasmid pCW3, is essential for efficient conjugative DNA transfer. Our studies reveal that TcpK is a member of the winged helix-turn-helix (wHTH) transcription factor superfamily and that it forms a dimer in solution. Furthermore, TcpK specifically binds to a nine-nucleotide sequence that is present as tandem repeats within the pCW3 origin of transfer (oriT). The X-ray crystal structure of the TcpK–TcpK box complex reveals a binding mode centered on and around the β-wing, which is different from what has been previously shown for other wHTH proteins. Structure-guided mutagenesis experiments validate the specific interaction between TcpK and the DNA molecule. Additional studies highlight that the TcpK dimer is important for specific DNA binding.

AB - Conjugation is fundamental for the acquisition of new genetic traits and the development of antibiotic resistance in pathogenic organisms. Here, we show that a hypothetical Clostridium perfringens protein, TcpK, which is encoded by the tetracycline resistance plasmid pCW3, is essential for efficient conjugative DNA transfer. Our studies reveal that TcpK is a member of the winged helix-turn-helix (wHTH) transcription factor superfamily and that it forms a dimer in solution. Furthermore, TcpK specifically binds to a nine-nucleotide sequence that is present as tandem repeats within the pCW3 origin of transfer (oriT). The X-ray crystal structure of the TcpK–TcpK box complex reveals a binding mode centered on and around the β-wing, which is different from what has been previously shown for other wHTH proteins. Structure-guided mutagenesis experiments validate the specific interaction between TcpK and the DNA molecule. Additional studies highlight that the TcpK dimer is important for specific DNA binding.

KW - bacterial genetics

KW - DNA

KW - x-ray crystallography

UR - http://www.scopus.com/inward/record.url?scp=85053339433&partnerID=8YFLogxK

U2 - 10.1038/s41467-018-06096-2

DO - 10.1038/s41467-018-06096-2

M3 - Article

VL - 9

JO - Nature Communications

JF - Nature Communications

SN - 2041-1723

IS - 1

M1 - 3732

ER -