S R Buchstein | PubMed Health

PubMed1994

Complete nucleotide sequence of a circular plasmid from the Lyme disease spirochete, Borrelia burgdorferi.

We have determined the complete nucleotide sequence of a small circular plasmid from the spirochete Borrelia burgdorferi Ip21, the agent of Lyme disease. The plasmid (cp8.3/Ip21) is 8,303 bp long, has a 76.6% A+T content, and is unstable upon passage of cells in vitro. An analysis of the sequence revealed the presence of two nearly perfect copies of a 184-bp inverted repeat sequence separated by 2,675 bp containing three closely spaced, but nonoverlapping, open reading frames (ORFs). Each inverted repeat ends in sequences that may function as signals for the initiation of transcription and translation of flanking plasmid sequences. A unique oligonucleotide probe based on the repeated sequence showed that the DNA between the repeats is present predominantly in a single orientation. Additional copies of the repeat were not detected elsewhere in the Ip21 genome. An analysis for potential ORFs indicates that the plasmid has nine highly probable protein-coding ORFs and one that is less probable; together, they occupy almost 71% of the nucleotide sequence. Analysis of the deduced amino acid sequences of the ORFs revealed one (ORF-9) with features in common with Borrelia lipoproteins and another (ORF-2) having limited homology with a replication protein, RepC, from a gram-positive plasmid that replicates by a rolling circle (RC) mechanism. Known collectively as RC plasmids, such plasmids require a double-stranded origin at which the Rep protein nicks the DNA to generate a single-stranded replication intermediate. cp8.3/Ip21 has three copies of the heptameric motif characteristically found at a nick site of most RC plasmids. These observations suggest that cp8.3/Ip21 may replicate by an RC mechanism.

Amino Acid Sequence↗

PubMed1991

Lyme disease.

The rapid rate of increase in reported cases of Lyme disease exceeds that of any zoonotic disease, and the annual occurrence of cases is greater than the sum of all other tick-borne illnesses in the United States. This, in addition to the geographic spread of cases to most of the United States, has greatly heightened investigative interests in all aspects of Lyme disease and its control.

Animals↗

PubMed1982

Genetic analysis of two bacterial RNA polymerase mutants that inhibit the growth of bacteriophage T7.

The Escherichia coli mutants 7009 and BR3 are defective in the growth of bacteriophage T7. We have previously shown that both of these mutant hosts produce an altered RNA polymerase which is resistant to inhibition by the T7 gene 2 protein (De Wyngaert and Hinkle 1979). In both strains, the mutation which prevents T7 growth is closely linked to rifA (rpoB). Both mutants are complemented by transformation with a multicopy plasmid carrying rpoB and rpoC but not by a plasmid carrying only rpoB. This indicates that the mutations reside in rpoC, the structural gene for the beta' subunit of RNA polymerase. When a single copy of the wildtype rpoC allele is introduced into the mutant using the transducing phage lambda drifd18, the mutant allele is dominant over wildtype. The lambda drifd18 transductant also remains unable to support the growth of T7 in the presence of rifampin. This supports our conclusion that the mutation is in rpoC. We have measured the growth of T7 phage, the kinetics of phage DNA synthesis, and the structure of replicative DNA intermediates in several transductants, and compared these results with those obtained in the original mutant strains.

DNA↗