Biomedical subjects

E C Sisk

Publications and source records attributed to E C Sisk.

5 recordsLinked to original sources

PubMed1999

Complete sequence of a 184-kilobase catabolic plasmid from Sphingomonas aromaticivorans F199.

The complete 184,457-bp sequence of the aromatic catabolic plasmid, pNL1, from Sphingomonas aromaticivorans F199 has been determined. A total of 186 open reading frames (ORFs) are predicted to encode proteins, of which 79 are likely directly associated with catabolism or transport of aromatic compounds. Genes that encode enzymes associated with the degradation of biphenyl, naphthalene, m-xylene, and p-cresol are predicted to be distributed among 15 gene clusters. The unusual coclustering of genes associated with different pathways appears to have evolved in response to similarities in biochemical mechanisms required for the degradation of intermediates in different pathways. A putative efflux pump and several hypothetical membrane-associated proteins were identified and predicted to be involved in the transport of aromatic compounds and/or intermediates in catabolism across the cell wall. Several genes associated with integration and recombination, including two group II intron-associated maturases, were identified in the replication region, suggesting that pNL1 is able to undergo integration and excision events with the chromosome and/or other portions of the plasmid. Conjugative transfer of pNL1 to another Sphingomonas sp. was demonstrated, and genes associated with this function were found in two large clusters. Approximately one-third of the ORFs (59 of them) have no obvious homology to known genes.

Bacterial Proteins↗

PubMed1998

Identification and sequence analysis of a 27-kilobase chromosomal fragment containing a Salmonella pathogenicity island located at 92 minutes on the chromosome map of Salmonella enterica serovar typhimurium LT2.

Using a genomic approach, we have identified a new Salmonella pathogenicity island, SPI-4, which is the fourth Salmonella pathogenicity island to be identified. SPI-4 was located at 92 min on the chromosome map and was flanked by the ssb and soxSR loci. The DNA sequence covering the entire SPI-4 and both boundaries was determined. The size of SPI-4 was about 25 kb and it contains 18 putative open reading frames (ORFs). Three of these ORFs encode proteins that have significant homology with proteins involved in toxin secretion. Another five ORFs encode proteins that have significant homology with hypothetical proteins from Synechocystis sp. strain PCC6803 or Acinetobacter calcoaceticus. The rest of the ORFs encode novel proteins, one of which has five membrane-spanning domains. SPI-4 is likely to carry a type I secretion system involved in toxin secretion. Furthermore, a previously identified locus (ims98), which is required for intramacrophage survival, was also mapped within the SPI-4 region. These findings suggested that SPI-4 is needed for intramacrophage survival.

Amino Acid Sequence↗

PubMed1995

Induction of base damage in DNA solutions by ultrasonic cavitation.

Ultrasound can damage macromolecules by the mechanical (shearing) and sonochemical (free radical generating) action of ultrasonic cavitation. Attributing macromolecular damage to either direct mechanical stress or to indirect mechanisms involving free radicals or other sonochemicals is a challenging problem. DNA damage induced by ultrasound was evaluated by measuring the formation of purine and pyrimidine products using combined gas chromatography-mass spectrometry with selected ion monitoring. Samples of DNA were prepared in 10 mmol dm-3 phosphate buffered saline (pH 7.4) and saturated with a mixture of argon:oxygen (3:1). Continuous 2.17 MHz ultrasound exposures at 0.82 mPa spatial peak negative pressure amplitude were performed in a 60 rpm rotating tube exposure system. Hydrogen peroxide yields were measured after each exposure to quantify the cavitation activity and ranged up to 350 mumol dm-3 for 1-h exposures. Purine and pyrimidine products identified were those typically observed following exposure of DNA to hydroxyl radical-generating systems, such as ionizing radiation, hypoxanthine/xanthine oxidase, or hydrogen peroxide in the presence of transition metal ions. The yields of these products were directly correlated with cavitation activity as measured by residual hydrogen peroxide concentrations. The yields of DNA products increased in the following order: thymine glycol approximately cytosine glycol > 8-oxoAde > FAPyAde approximately 5-HMU approximately 5,6-diOHCyt > FAPyGua. Unexpectedly, 8-oxoguanine did not exhibit a dose-dependent increase above background levels, and this observation is inconsistent with processes involving metal ion-dependent formation of hydroxyl radicals from hydrogen peroxide. In addition, the product yields were far too large to result from the residual hydrogen peroxide. Thus, ultrasonic cavitation appears to have a mode of action distinct from either ionizing radiation or formation of hydroxyl radicals via Fenton-like reaction with transition metals.

Adenine↗

PubMed1994

Electron migration in oligonucleotides upon gamma-irradiation in solution.

Electron migration in irradiated solutions of DNA was investigated using 5-bromouracil synthetically incorporated into oligonucleotides of defined base composition as a molecular indicator of electron interactions. Solvated electrons interact quantitatively with 5-bromouracil, leading to a highly reactive 5-yl radical which can abstract an adjacent hydrogen atom to yield uracil. Yields of uracil, or loss of 5-bromouracil, from irradiated oligonucleotide samples were measured using gas chromatography-mass spectrometric analysis of their trimethylsilylated acid hydrolysates. To examine the effects of base composition and DNA conformation on electron migration, a set of oligonucleotides containing 5-bromouracil at selected positions with three base (guanine, cytosine, thymine or adenine) spacers (e.g. [BrU(GGG)3]3) were irradiated in their single- or double-stranded form following annealing with appropriate complementary sequences. Differences in uracil yields suggested that electron migration occurred to different extents in oligonucleotides containing different base sequences. In irradiated single-stranded oligonucleotides, the yield of uracil decreased in the order A > T > > C approximately G. However, in irradiated double-stranded oligonucleotides, the yield of uracil decreased in the order G > C approximately T > A. These differences were attributed to proton-transfer reactions facilitated by base pairing in double-stranded oligonucleotides. The distance over which the electron would migrate was then determined using a series of oligonucleotides containing 5-bromouracil at selected positions with guanine spacers (i.e. [BrU(G)n]3 (n = 3, 5, 7, 9). Oligonucleotides were irradiated in their double-stranded form following annealing with the appropriate complementary sequences. Analysis of the loss of 5-bromouracil revealed that electron migration occurred efficiently over c. 3-4 guanine bases assuming that migration could occur as efficiently in either direction along the DNA molecule. These data can be compared with studies reporting more extensive migration for electrons generated by direct ionization of DNA.

Base Sequence↗

PubMed1994

Radiation-induced electron migration in nucleic acids.

Radiation-induced electron migration along DNA is a mechanism by which randomly produced stochastic energy deposition events can lead to non-random types of damage along DNA manifested distal to the sites of the initial energy deposition. Radiation-induced electron migration in nucleic acids has been examined using oligonucleotides containing 5-bromouracil (5-BrU). Interaction of 5-BrU with solvated electrons results in release of bromide ions and formation or uracil-5-yl radicals. Monitoring either bromide ion release or uracil formation provides an opportunity to study electron migration processes in model nucleic acid systems. Using this approach we have discovered that electron migration along oligonucleotides is significantly influenced by the base sequence and strandedness. Migration along 7 base pairs in oligonucleotides containing guanine bases was observed for oligonucleotides irradiated in solution, which compares with mean migration distances of 6-10 bp for Escherichia coli DNA irradiated in solution and 5.5 bp for E. coli DNA irradiated in cells. Evidence also suggests that electron migration can occur preferentially in the 5' to 3' direction along a double-stranded oligonucleotide containing a region of purine bases adjacent to the 5-BrU moiety. Our continued efforts will provide information regarding the contribution of electron transfer along DNA to formation of locally multiply damaged sites created in DNA by exposure to ionizing radiation.

Base Sequence↗