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DNA Microarray Analysis of Genome Dynamics in Yersinia pestis: Insights into Bacterial Genome Microevolution and Niche Adaptation. Dongsheng Zhou, 2004.Genomics research provides an unprecedented opportunity for us to probe into the pathogenicity and evolution of the world's most deadly pathogenic bacterium, Yersinia pestis, in minute detail . In our present work, extensive microarray analysis in conjunction with PCR validation revealed that there are considerable genome dynamics, due to gene acquisition and loss, in natural populations of Y . pestis . We established a genomotyping system to group homologous isolates of Y . pestis, based on profiling or gene acquisition and loss in their genomes, and then drew an outline of parallel microevolution of the Y . pestis genome . The acquisition of a number of genomic islands and plasmids most likely induced Y . pestis to evolve rapidly from Yersinia pseudotuberculosis to a new, deadly pathogen . Horizontal gene acquisition also plays a key role in the dramatic evolutionary segregation of Y . pestis lineages (biovars and genomovars) . In contrast to selective genome expansion by gene acquisition, genome reduction occurs in Y . pestis through the loss of DNA regions . We also theorized about the links between niche adaptation and genome microevolution . The transmission, colonization, and expansion of Y . pestis in the natural foci of endemic plague are parallel and directional and involve gradual adaptation to the complex of interactions between the environment, the hosts, and the pathogen itself . These adaptations are based on the natural selections against the accumulation of genetic changes within genome . Our data strongly support that the modern plague originated from Yunnan Province in China, due to the arising of biovar orientalis from biovar antiqua rather than mediaevalis . Regulation of the Alternative Sigma Factor  E during Initiation, Adaptation, and Shutoff of the Extracytoplasmic Heat Shock Response in Escherichia coli.Sarah E. Ades, 2003.The alternative sigma factor E is activated in response to stress in the extracytoplasmic compartment of Escherichia coli . Here we show that
E activity increases upon initiation of the stress response by a shift to an elevated temperature (43°C) and remains at that level for the duration of the stress . When the stress is removed by a temperature downshift,
E activity is strongly repressed and then slowly returns to levels seen in unstressed cells . We provide evidence that information about the state of the cell envelope is communicated to
E primarily through the regulated proteolysis of the inner membrane anti-sigma factor RseA, as the degradation rate of RseA is correlated with the changes in
E activity throughout the stress response . However, the relationship between
E activity and the rate of degradation of RseA is complex, indicating that other factors may cooperate with RseA and serve to fine-tune the response .
Evolutionary Engineering of Saccharomyces cerevisiae for Anaerobic Growth on Xylose. Marco Sonderegger, 2003.Xylose utilization is of commercial interest for efficient conversion of abundant plant material to ethanol . Perhaps the most important ethanol-producing organism, Saccharomyces cerevisiae, however, is incapable of xylose utilization . While S . cerevisiae strains have been metabolically engineered to utilize xylose, none of the recombinant strains or any other naturally occurring yeast has been able to grow anaerobically on xylose . Starting with the recombinant S . cerevisiae strain TMB3001 that overexpresses the xylose utilization pathway from Pichia stipitis, in this study we developed a selection procedure for the evolution of strains that are capable of anaerobic growth on xylose alone . Selection was successful only when organisms were first selected for efficient aerobic growth on xylose alone and then slowly adapted to microaerobic conditions and finally anaerobic conditions, which indicated that multiple mutations were necessary . After a total of 460 generations or 266 days of selection, the culture reproduced stably under anaerobic conditions on xylose and consisted primarily of two subpopulations with distinct phenotypes . Clones in the larger subpopulation grew anaerobically on xylose and utilized both xylose and glucose simultaneously in batch culture, but they exhibited impaired growth on glucose . Surprisingly, clones in the smaller subpopulation were incapable of anaerobic growth on xylose . However, as a consequence of their improved xylose catabolism, these clones produced up to 19% more ethanol than the parental TMB3001 strain produced under process-like conditions from a mixture of glucose and xylose . CynD, the Cyanide Dihydratase from Bacillus pumilus: Gene Cloning and Structural Studies. Dakshina Jandhyala, 2003.The cyanide dihydratase in Bacillus pumilus was shown to be an 18-subunit spiral structure by three-dimensional reconstruction of electron micrographs of negatively stained material at its optimum pH, 8.0 . At pH 5.4, the subunits rearrange to form an extended left-handed helix . Gel electrophoresis of glutaraldehyde cross-linked enzyme suggests that the fundamental component of the spiral is a dimer of the 37-kDa subunit . The gene was cloned, and the recombinant enzyme was readily expressed at high levels in Escherichia coli . Purification of the recombinant enzyme was facilitated by the addition of a C-terminal six-histidine affinity purification tag . The tagged recombinant enzyme has Km and Vmax values similar to those published for the native enzyme . This is the first cyanide dihydratase from a gram-positive bacterium to be sequenced, and it is the first description of the structure of any member of this enzyme class . The putative amino acid sequence shares over 80% identity to the only other sequenced cyanide dihydratase, that of the gram-negative Pseudomonas stutzeri strain AK61, and is similar to a number of other bacterial and fungal nitrilases . This sequence similarity suggests that the novel short spiral structure may be typical of these enzymes . In addition, an active cyanide dihydratase from a non-cyanide-degrading isolate of B . pumilus (strain 8A3) was cloned and expressed . This suggests that cynD, the gene coding for the cyanide dihydratase, is not unique to the C1 strain of B . pumilus and is not a reflection of its origin at a mining waste site .
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