Dataset Information

Phage inhibition, colicin resistance, and tellurite resistance are encoded by a single cluster of genes on the IncHI2 plasmid R478.

ABSTRACT: A region of the IncHI2 plasmid R478, encoding the phenotypes of tellurite resistance (Ter), phage inhibition (Phi), and colicin resistance (PacB), was cloned and sequenced. Analysis indicated seven open reading frames (ORFs), whose genes were designated terZ, -A, -B, -C, -D, -E, and -F. Five of these predicted ORFs (A to E) had extensive amino acid homology with the previously reported ORFs of the IncHI2 Ter operon from plasmid pMER610. There were domains of highly conserved amino acid residues within the group TerA, -D, -E, and -F and within TerD, -E, and -Z, but no consensus could be found among all five putative polypeptides. There were also regions of good identity and similarity between individual pairs of ORFs which was not reflected in the multiple alignments. The three phenotypes were expressed in Escherichia coli DH5 alpha by an 8.4-kb EcoRI insert subcloned from a cosmid of R478. The latter insert was clonable only as a double insertion with a 4.5-kb fragment, and forced deletion of the smaller fragment was lethal to cells. This lethality was not dependent on the cloned orientation of either fragment, suggesting that there is a trans-acting element in the 4.5-kb fragment. Tn1000 mutagenesis of one of the double-insert clones, pDT2575, showed that the phenotypes, including multiple colicin resistance, were genetically linked. Transpositions into terD, terC, and terZ reduced or abolished all phenotypes, while inserts into terE and terF had no effect on the phenotypes. Insertions in terA reduced phage inhibition levels only. The presence of the terZ and terF ORFs in pMER610 was confirmed, and derivatives of this plasmid mediated Phi, PacB, and Ter.

SUBMITTER: Whelan KF

PROVIDER: S-EPMC177279 | biostudies-other | 1995 Sep

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

Phage inhibition, colicin resistance, and tellurite resistance are encoded by a single cluster of genes on the IncHI2 plasmid R478.

Whelan K F KF Colleran E E Taylor D E DE

Journal of bacteriology 19950901 17

A region of the IncHI2 plasmid R478, encoding the phenotypes of tellurite resistance (Ter), phage inhibition (Phi), and colicin resistance (PacB), was cloned and sequenced. Analysis indicated seven open reading frames (ORFs), whose genes were designated terZ, -A, -B, -C, -D, -E, and -F. Five of these predicted ORFs (A to E) had extensive amino acid homology with the previously reported ORFs of the IncHI2 Ter operon from plasmid pMER610. There were domains of highly conserved amino acid residues ...[more]

PMID: 7665479

Similar Datasets

Project description:Bacteriophages (phages) and other viruses are extremely efficient in packing their genetic information, with several described cases of overlapping genes encoded in different open reading frames (ORFs). While less frequently reported, specific cases exist in which two overlapping ORFs are in frame and share the stop codon. Here, we studied the occurrence of this genetic arrangement in endolysins, the phage enzymes that cut the bacterial cell wall peptidoglycan to release the virion progeny. After screening over 3,000 endolysin sequences of phages infecting Gram-positive bacteria, we found evidence that this coding strategy is frequent in endolysin genes. Our bioinformatics predictions were experimentally validated by demonstrating that two polypeptides are indeed produced from these genes. Additionally, we show that in some cases the two polypeptides need to interact and multimerize to generate the active endolysin. By studying in detail one selected example, we uncovered a heteromeric endolysin with a 1:5 subunit stoichiometry that has never been described before. Hence, we conclude that the occurrence of endolysin genes encoding two polypeptide isoforms by in-frame overlapping ORFs, as well as their organization as enzymatic complexes, appears more common than previously thought, therefore challenging the established view of endolysins being mostly formed by single, monomeric polypeptide chains. IMPORTANCE Bacteriophages use endolysins to cleave the host bacteria cell wall, a crucial event underlying cell lysis for virion progeny release. These bacteriolytic enzymes are generally thought to work as single, monomeric polypeptides, but a few examples have been described in which a single gene produces two endolysin isoforms. These are encoded by two in-frame overlapping ORFs, with a shorter ORF being defined by an internal translation start site. This work shows evidence that this endolysin coding strategy is frequent in phages infecting Gram-positive bacteria, and not just an eccentricity of a few phages. In one example studied in detail, we show that the two isoforms are inactive until they assemble to generate a multimeric active endolysin, with a 1:5 subunit stoichiometry never described before. This study challenges the established view of endolysins, with possible implications in their current exploration and design as alternative antibacterials.

Dataset Information

Phage inhibition, colicin resistance, and tellurite resistance are encoded by a single cluster of genes on the IncHI2 plasmid R478.

Publications

Phage inhibition, colicin resistance, and tellurite resistance are encoded by a single cluster of genes on the IncHI2 plasmid R478.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets