Biomedical subjects

Yoshio Kimura

Publications and source records attributed to Yoshio Kimura.

9 recordsLinked to original sources

PubMed2006

Contribution of the cyclic nucleotide phosphodiesterases PdeA and PdeB to adaptation of Myxococcus xanthus cells to osmotic or high-temperature stress.

A tBLASTn search of the Myxococcus xanthus genome database at The Institute for Genomic Research (TIGR) identified three genes (pdeA, pdeB, and pdeC) that encode proteins homologous to 3',5'-cyclic nucleotide phosphodiesterase. pdeA, pdeB, and pdeC mutants, constructed by replacing a part of the gene with the kanamycin or tetracycline resistance gene, showed normal growth, development, and germination under nonstress conditions. However, the spores of mutants, especially the pdeA and pdeB mutants, placed under osmotic stress germinated earlier than the wild-type spores. The phenotype was the opposite of that of the receptor-type adenylyl cyclase (cyaA or cyaB) mutant. Also, pdeA and pdeB mutants were found to have impaired growth under the condition of high-temperature stress. Intracellular cyclic AMP (cAMP) levels of pdeA or pdeB mutant cells under these stressful conditions were about 1.3-fold to 2.0-fold higher than those of wild-type cells. These results suggest that PdeA and PdeB may be involved in osmotic adaptation during spore germination and temperature adaptation during vegetative growth through the regulation of cAMP levels.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed2005

Myxococcus xanthus twin-arginine translocation system is important for growth and development.

The twin-arginine translocation (Tat) system serves to export fully folded proteins across the cytoplasmic membrane. In many bacteria, three major components, TatA, TatB and TatC, are the functionally essential constituents of the Tat system. A Myxococcus xanthus tatB-tatC deletion mutant could aggregate and form mounds, but was unable to form fruiting bodies under nutritionally limiting conditions. When tatB-tatC mutant vegetative cells were cultured with 0.5 M glycerol, the cell morphology changed to spore-like spherical cells, but the spores were not resistant to heat and sonication treatments. In contrast to the wild-type strain, the tatB-tatC mutant also showed a decreased cell growth rate and a lower maximum cell concentration. These results suggest possibility that the Tat system may contribute to export of various important proteins for development and growth for M. xanthus.

Amino Acid Sequence↗

PubMed2005

A Myxococcus xanthus CbpB containing two cAMP-binding domains is involved in temperature and osmotic tolerances.

Our previous data indicated that a Myxococcus xanthus sensor-type adenylyl cyclase (CyaA) functions in signal transduction during osmotic stress. However, the cAMP-mediated signal transduction pathway in this bacterium was unknown. Here, we isolated a clone from a M. xanthus genomic DNA library using oligonucleotide probes designed based on the conserved cAMP-binding domains of the cAMP-dependent protein kinase (PKA) regulatory subunits. The clone contained two open-reading frames (ORFs), cbpA and cbpB, encoding hydrophilic proteins with one and two cAMP-binding domains, respectively. The CbpB exhibited partial primary structural similarity to PKA regulatory subunits. cbpA and cbpB mutants, generated by gene disruption, showed normal growth, development and spore germination. However, the cbpB mutant cultured under high- or low-temperature conditions exhibited a marked reduction in growth. cbpB mutant cells were also more sensitive to osmotic stress than wild-type cells. The cbpA mutant possessed normal resistance to such stress. The phenotype of cbpB mutant was similar to those of PKA regulatory subunit mutants of some eukaryotic microorganisms.

Amino Acid Sequence↗

PubMed2005

An adenylyl cyclase, CyaB, acts as an osmosensor in Myxococcus xanthus.

We have previously reported that a receptor-type adenylyl cyclase (CyaA) of Myxococcus xanthus undergoes an osmosensor mainly during spore germination (Y. Kimura et al., J. Bacteriol. 184:3578-3585, 2002). In the present study, we cloned another receptor-type adenylyl cyclase gene (cyaB) and characterized the function of the cyaB-encoded protein. Disruption of cyaB generates a mutant that showed growth retardation at high ionic (NaCl) or high nonionic (sucrose) osmolarity. When vegetative cells were stimulated with 0.15 M NaCl, the increases in intracellular cyclic AMP levels of cyaB mutant cells were lower than those of wild-type cells. Under nonionic osmostress, the cyaB mutant exhibited reduced spore germination; however, the germination rate of the cyaB mutant was significantly higher than that of the cyaA mutant.

Adenylyl Cyclases↗

PubMed2004

A Myxococcus xanthus rppA-mmrA double mutant exhibits reduced uptake of amino acids and tolerance of some antimicrobials.

Myxococcus xanthus RppA and MmrA are homologous to methyl-accepting chemotaxis proteins (MCPs) and to multidrug transporters, respectively. We reported previously that rppA-mmrA double mutant exhibited reduced colony expansion, agglutination, and polysaccharide levels. We have demonstrated here that the rppA-mmrA mutant also exhibited reduced amino acid uptake. Furthermore, the double mutant appeared to be more susceptible to some antimicrobial agents, such as streptomycin, ethidium bromide and norfloxacin, than the wild-type. These phenotypes were not shown in the rppA or mmrA single mutant. These results indicate that M. xanthus RppA and MmrA are also involved in the uptake of amino acids and efflux of some antimicrobial agents.

Anti-Bacterial Agents↗

PubMed2004

RppA, a transducer homologue, and MmrA, a multidrug transporter homologue, are involved in the biogenesis and/or assembly of polysaccharide in Myxococcus xanthus.

Myxococcus xanthus cells move by gliding, and form multicellular fruiting bodies under conditions of starvation. The authors cloned a gene, designated rppA (for receptor for polysaccharide production), which encodes a methyl-accepting protein homologous to the chemotaxis transducers in eubacteria. The rppA gene was co-transcribed with mmrA, a gene homologous to various multidrug transporter genes. The rppA or mmrA single mutants showed almost identical phenotypes to the wild-type strain; however, the rppA-mmrA double mutant exhibited reduced colony expansion, cell-cell agglutination and cellular reversal frequency. The double-mutant cells also showed less binding to Congo red, which mainly binds to fibril polysaccharide, than wild-type cells. Analysis of total polysaccharide in stationary-phase cells demonstrated that in the double mutant, polysaccharide levels were decreased by about 30 % as compared with the wild-type strain. These results indicated that RppA and MmrA play a role in the biogenesis and/or assembly of polysaccharide, and the phenotypes of the double mutant may be due to the reduction in fibril polysaccharide.

ATP Binding Cassette Transporter, Subfamily B↗

PubMed2003

[Precision of internal standard method in HPLC analysis].

The internal standard methods are known to compensate for the errors from sample preparation and injection into an analytical instrument. However, recent HPLC apparatuses have injectors of excellent repeatability and it is dubious whether the cancellation of injection error can lead to substantial improvement in the precision of analysis. This paper answers the above question experimentally and theoretically. The HPLC analysis of butylscopolamine bromide is taken as an example. The relative standard deviations (RSD) of measurements in the internal standard method and absolute calibration curve method are compared and the advantages of these methods are discussed. The measurement RSD is shown to be well estimated by the (function of mutual information) (FUMI) theory without repeating measurements. This report also demonstrates simple equations for calculating the measurement RSD at an arbitrary concentration of analyte and for selecting the better method between the internal standard method and absolute calibration curve method under specific experimental conditions.

Calibration↗

PubMed2002

An adenylyl cyclase, CyaA, of Myxococcus xanthus functions in signal transduction during osmotic stress.

An adenylyl cyclase gene (cyaA) present upstream of an osmosensor protein gene (mokA) was isolated from Myxococcus xanthus. cyaA encoded a polypeptide of 843 amino acids with a predicted molecular mass of 91,187 Da. The predicted cyaA gene product had structural similarity to the receptor-type adenylyl cyclases that are composed of an amino-terminal sensor domain and a carboxy-terminal catalytic domain of adenylyl cyclase. In reverse transcriptase PCR experiments, the transcript of the cyaA gene was detected mainly during development and spore germination. A cyaA mutant, generated by gene disruption, showed normal growth, development, and germination. However, a cyaA mutant placed under conditions of ionic (NaCl) or nonionic (sucrose) osmostress exhibited a marked reduction in spore formation and spore germination. When wild-type and cyaA mutant cells at developmental stages were stimulated with 0.2 M NaCl or sucrose, the mutant cells increased cyclic AMP accumulation at levels similar to those of the wild-type cells. In contrast, the mutant cells during spore germination had mainly lost the ability to respond to high-ionic osmolarity. In vegetative cells, the cyaA mutant responded normally to osmotic stress. These results suggested that M. xanthus CyaA functions mainly as an ionic osmosensor during spore germination and that CyaA is also required for osmotic tolerance in fruiting formation and sporulation.

Adenylate Cyclase Toxin↗

PubMed2002

[Estimation of precision in system suitability tests].

In system suitability tests, the daily conditions of an analytical instrument are checked with the standard deviation (SD) of measurements as a criterion. This paper examines how exactly the SD values obtained from the measurements can indicate instrumental conditions. The HPLC measurement for acetaminophen is repeated six times daily to obtain an SD estimate, and these procedures are repeated for seven days. The degree of scattering of the seven SD values corresponds well to the 95% confidence intervals calculated from the chi-square distribution (n = 6). Therefore it is concluded that the variability of the daily conditions of the HPLC system used in this study is too small to detect by referring to the SD estimates (n = 6) and the daily change in the SD estimates only represents the error of the SD estimation itself. However, if the repetition number is 40 (n = 40), the SD estimates will be a good indicator. Forty measurements are practically impossible for a slow analysis like HPLC, and this paper recommends the function of mutual information theory as a substitute for repetitive measurement.

Acetaminophen↗