Regional specificity of illegitimate recombination associated with the translocatable ampicillin-resistance element Tn1.
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Biomedical subjects
Publications and source records attributed to G M Weinstock.
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Enterococci are a long-known cause of bacterial endocarditis and a more recently recognized cause of nosocomial infection and superinfection. While much is known about the many antibiotic resistances of enterococci, less is known about the organism itself and how it causes disease. This article presents a brief overview of enterococci and its possible virulence factors and summarizes the authors' efforts to understand the features of this organism that may contribute to its disease potential.
Each of the three cysteine residues in the Escherichia coli RecA protein was replaced with a number of other amino acids. To do this, each cysteine codon was first converted to a chain-terminating amber codon by oligonucleotide-directed mutagenesis. These amber mutants were then either assayed for function in different suppressor strains or reverted by a second round of mutagenesis with oligonucleotides that had random sequences at the amber codon. Thirty-three different amino acid substitutions were obtained. Mutants were tested for three functions of RecA: survival following UV irradiation, homologous recombination, and induction of the SOS response. It was found that although none of the cysteines is essential for activity, mutations at each of these positions can affect one or more of the activities of RecA, depending on the particular amino acid substitution. In addition, the cysteine at position 116 appears to be involved in the RecA-promoted cleavage of the LexA protein.
Bovine serum was used to identify a recombinant phage clone carrying the Pasteurella haemolytica leukotoxin gene. This fragment produced the 102-kD leukotoxin and several smaller P. haemolytica-specific protein antigens in Escherichia coli. An additional contiguous fragment, containing sequences upstream from the leukotoxin gene. Using these clones, we determined the nucleotide sequence of a 7745-bp region that included four open reading frames: an upstream gene, lktC; the leukotoxin gene, lktA; and two downstream genes, lktB, and lktD. The predicted molecular weights of the proteins encoded by these genes were 19.9, 102, 79.6, and 54.7 kD, respectively. These genes and their predicted proteins were similar in organization and in sequence to the corresponding elements of the gene cluster that encodes an E. coli alpha-hemolysin and its activation and secretion functions. Expression of the leukotoxin was enhanced in E. coli, by fusing the gene to the lac promoter. Under these conditions the leukotoxin was not secreted into the medium, as it is in P. haemolytica. However, in the presence of the alpha-hemolysin genes, the leukotoxin was secreted into the medium, demonstrating functional complementation by the hemolysin secretory system.
Whole-genome sequencing is transforming the study of pathogenic bacteria. Searches for single virulence genes can now be performed on a genomewide scale by a variety of computer and genetic techniques. These techniques are discussed to provide a perspective on the developing field of genomics.