Dataset Information

CDP-alcohol hydrolase, a very efficient activity of the 5'-nucleotidase/UDP-sugar hydrolase encoded by the ushA gene of Yersinia intermedia and Escherichia coli.

ABSTRACT: Nucleoside 5'-diphosphate-X hydrolases are interesting enzymes to study due to their varied activities and structure-function relationships and the roles they play in the disposal, assimilation, and modulation of the effects of their substrates. Few of these enzymes with a preference for CDP-alcohols are known. In Yersinia intermedia suspensions prepared from cultures on Columbia agar with 5% sheep blood, we found a CDP-alcohol hydrolase liberated to Triton X-100-containing medium. Growth at 25 degrees C was deemed optimum in terms of the enzyme-activity yield. The purified enzyme also displayed 5'-nucleotidase, UDP-sugar hydrolase, and dinucleoside-polyphosphate hydrolase activities. It was identified as the protein product (UshA(Yi)) of the Y. intermedia ushA gene (ushA(Yi)) by its peptide mass fingerprint and by PCR cloning and expression to yield active enzyme. All those activities, except CDP-alcohol hydrolase, have been shown to be the properties of UshA of Escherichia coli (UshA(Ec)). Therefore, UshA(Ec) was expressed from an appropriate plasmid and tested for CDP-alcohol hydrolase activity. UshA(Ec) and UshA(Yi) behaved similarly. Besides being the first study of a UshA enzyme in the genus Yersinia, this work adds CDP-alcohol hydrolase to the spectrum of UshA activities and offers a novel perspective on these proteins, which are viewed here for the first time as highly efficient enzymes with k(cat)/K(m) ratios near the theoretical maximum level of catalytic activities. The results are discussed in the light of the known structures of UshA(Ec) conformers and the respective homology models constructed for UshA(Yi), and also in relation to possible biological functions. Interestingly, every Yersinia species with a sequenced genome contains an intact ushA gene, except Y. pestis, which in all its sequenced biovars contains a ushA gene inactivated by frameshift mutations.

SUBMITTER: Alves-Pereira I

PROVIDER: S-EPMC2546799 | biostudies-literature | 2008 Sep

REPOSITORIES: biostudies-literature

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CDP-alcohol hydrolase, a very efficient activity of the 5'-nucleotidase/UDP-sugar hydrolase encoded by the ushA gene of Yersinia intermedia and Escherichia coli.

Alves-Pereira Isabel I Canales José J Cabezas Alicia A Cordero Paloma Martín PM Costas María Jesús MJ Cameselle José Carlos JC

Journal of bacteriology 20080718 18

Nucleoside 5'-diphosphate-X hydrolases are interesting enzymes to study due to their varied activities and structure-function relationships and the roles they play in the disposal, assimilation, and modulation of the effects of their substrates. Few of these enzymes with a preference for CDP-alcohols are known. In Yersinia intermedia suspensions prepared from cultures on Columbia agar with 5% sheep blood, we found a CDP-alcohol hydrolase liberated to Triton X-100-containing medium. Growth at 25 ...[more]

PMID: 18641143

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Project description:BackgroundThe yersiniae (Enterobacteriaceae) occupy a variety of niches, including some in human and flea hosts. Metabolic adaptations of the yersiniae, which contribute to their success in these specialized environments, remain largely unknown. We report results of an investigation of the transcriptome under aerobic and anaerobic conditions for Y. intermedia, a non-pathogenic member of the genus that has been used as a research surrogate for Y. pestis. Y. intermedia shares characteristics of pathogenic yersiniae, but is not known to cause disease in humans. Oxygen restriction is an important environmental stimulus experienced by many bacteria during their life-cycles and greatly influences their survival in specific environments. How oxygen availability affects physiology in the yersiniae is of importance in their life cycles but has not been extensively characterized.Methodology/principal findingsTiled oligonucleotide arrays based on a draft genome sequence of Y. intermedia were used in transcript profiling experiments to identify genes that change expression in response to oxygen availability during growth in minimal media with glucose. The expression of more than 400 genes, constituting about 10% of the genome, was significantly altered due to oxygen-limitation in early log phase under these conditions. Broad functional categorization indicated that, in addition to genes involved in central metabolism, genes involved in adaptation to stress and genes likely involved with host interactions were affected by oxygen-availability. Notable among these, were genes encoding functions for motility, chemotaxis and biosynthesis of cobalamin, which were up-regulated and those for iron/heme utilization, methionine metabolism and urease, which were down-regulated.Conclusions/significanceThis is the first transcriptome analysis of a non-pathogenic Yersinia spp. and one of few elucidating the global response to oxygen limitation for any of the yersiniae. Thus this study lays the foundation for further experimental characterization of oxygen-responsive genes and pathways in this ecologically diverse genus.

Dataset Information

CDP-alcohol hydrolase, a very efficient activity of the 5'-nucleotidase/UDP-sugar hydrolase encoded by the ushA gene of Yersinia intermedia and Escherichia coli.

Publications

CDP-alcohol hydrolase, a very efficient activity of the 5'-nucleotidase/UDP-sugar hydrolase encoded by the ushA gene of Yersinia intermedia and Escherichia coli.

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