Dataset Information

HicAB toxin-antitoxin complex from Escherichia coli: expression and crystallization.

ABSTRACT: Toxin-antitoxin (TA) systems are widespread in both bacteria and archaea, where they enable cells to adapt to environmental cues. TA systems play crucial roles in various cellular processes, such as programmed cell death, cell growth, persistence and virulence. Here, two distinct forms of the type II toxin-antitoxin complex HicAB were identified and characterized in Escherichia coli K-12, and both were successfully overexpressed and purified. The two proposed forms, HicABL and HicABS, differed in the presence or absence of a seven-amino-acid segment at the N-terminus in the antitoxin HicB. The short form HicABS readily crystallized under the conditions 0.1?M Tris-HCl pH 8.0, 20%(w/v) PEG 6000, 0.2?M ammonium sulfate. The HicABS crystal diffracted and data were collected to 2.5?Å resolution. The crystal belonged to space group I222 or I212121, with unit-cell parameters a = 67.04, b = 66.31, c = 120.78?Å. Matthews coefficient calculation suggested the presence of two molecules each of HicA and HicBS in the asymmetric unit, with a solvent content of 55.28% and a Matthews coefficient (VM) of 2.75?Å3?Da-1.

SUBMITTER: Yang J

PROVIDER: S-EPMC5619741 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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HicAB toxin-antitoxin complex from Escherichia coli: expression and crystallization.

Yang Jingsi J Xu Bingshuang B Gao Zengqiang Z Zhou Ke K Liu Peng P Dong Yuhui Y Zhang Jianjun J Liu Quansheng Q

Acta crystallographica. Section F, Structural biology communications 20170814 Pt 9

Toxin-antitoxin (TA) systems are widespread in both bacteria and archaea, where they enable cells to adapt to environmental cues. TA systems play crucial roles in various cellular processes, such as programmed cell death, cell growth, persistence and virulence. Here, two distinct forms of the type II toxin-antitoxin complex HicAB were identified and characterized in Escherichia coli K-12, and both were successfully overexpressed and purified. The two proposed forms, HicAB<sup>L</sup> and HicAB<s ...[more]

PMID: 28876228

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Project description:Type II toxin-antitoxin (TA) systems are classically composed of two genes that encode a toxic protein and a cognate antitoxin protein. Both genes are organized in an operon whose expression is autoregulated at the level of transcription by the antitoxin-toxin complex, which binds operator DNA through the antitoxin's DNA-binding domain. Here, we investigated the transcriptional regulation of a particular TA system located in the immunity region of a cryptic lambdoid prophage in the Escherichia coli O157:H7 EDL933 strain. This noncanonical paaA2-parE2 TA operon contains a third gene, paaR2, that encodes a transcriptional regulator that was previously shown to control expression of the TA. We provide direct evidence that the PaaR2 is a transcriptional regulator which shares functional similarities to the lambda CI repressor. Expression of the paaA2-parE2 TA operon is regulated by two other transcriptional regulators, YdaS and YdaT, encoded within the same region. We argue that YdaS and YdaT are analogous to lambda Cro and CII and that they do not constitute a TA system, as previously debated. We show that PaaR2 primarily represses the expression of YdaS and YdaT, which in turn controls the expression of paaR2-paaA2-parE2 operon. Overall, our results show that the paaA2-parE2 TA is embedded in an intricate lambdoid prophage-like regulation network. Using single-cell analysis, we observed that the entire locus exhibits bistability, which generates diversity of expression in the population. Moreover, we confirmed that paaA2-parE2 is addictive and propose that it could limit genomic rearrangements within the immunity region of the CP-933P cryptic prophage. IMPORTANCE Transcriptional regulation of bacterial toxin-antitoxin (TA) systems allows compensation of toxin and antitoxin proteins to maintain a neutral state and avoid cell intoxication unless TA genes are lost. Such models have been primarily studied in plasmids, but TAs are equally present in other mobile genetic elements, such as transposons and prophages. Here, we demonstrate that the expression of a TA system located in a lambdoid cryptic prophage is transcriptionally coupled to the prophage immunity region and relies on phage transcription factors. Moreover, competition between transcription factors results in bistable expression, which generates cell-to-cell heterogeneity in the population, but without, however, leading to any detectable phenotype, even in cells expressing the TA system. We show that despite the lack of protein sequence similarity, this locus retains major lambda prophage regulation features.

Dataset Information

HicAB toxin-antitoxin complex from Escherichia coli: expression and crystallization.

Publications

HicAB toxin-antitoxin complex from Escherichia coli: expression and crystallization.

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