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Kazuo Kobayashi

Publications and source records attributed to Kazuo Kobayashi.

At least 73 records · Page 4Linked to original sources
PubMed2003

Structural analysis of sphingophospholipids derived from Sphingobacterium spiritivorum, the type species of genus Sphingobacterium.

The unique feature of the genus Sphingobacterium is the presence of sphingophospholipids and ceramides, besides diacylglycerophospholipids. As major cellular lipid components, five kinds of sphingophospholipids were purified from Sphingobacterium spiritivorum ATCC 33861(T), the type species of genus Sphingobacterium. They were identified as ceramide phosphorylethanolamines (CerPE-1 and CerPE-2), ceramide phosphoryl-myo-inositols (CerPI-1 and CerPI-2), and ceramide phosphorylmannose (CerPM-1). The ceramide of CerPE-1, CerPI-1, and CerPM-1 was composed of 15-methylhexadecasphinganine (isoheptadeca sphinganine, iso-C17:0) and 13-methyltetradecanoic acid (isopentadecanoic acid, iso-C15:0), whereas that of CerPE-2 and CerPI-2 was composed of isoheptadeca sphinganine and 2-hydroxy-13-methyltetradecanoic acid (2-hydroxy isopentadecanoic acid, 2-OH iso-C15:0). These sphingophospholipids were also found in cellular lipids of Sphingobacterium multivorum ATCC 33613(T), Sphingobacterium mizutaii ATCC 33299(T), Sphingobacterium faecium IFO 15299(T), Sphingobacterium thalpophilum ATCC 43320(T), and Sphingobacterium antarcticum ATCC 51969(T). To our knowledge, the existence of CerPM-1 is a novel sphingophospholipid through eukaryotic and prokaryotic cells.

Ceramides↗
PubMed2003

A functional link between RuBisCO-like protein of Bacillus and photosynthetic RuBisCO.

The genomes of several nonphotosynthetic bacteria, such as Bacillus subtilis, and some Archaea include genes for proteins with sequence homology to the large subunit of ribulose bisphosphate carboxylase/oxygenase (RuBisCO). We found that such a RuBisCO-like protein (RLP) from B. subtilis catalyzed the 2,3-diketo-5-methylthiopentyl-1-phosphate enolase reaction in the methionine salvage pathway. A growth-defective mutant, in which the gene for this RLP had been disrupted, was rescued by the gene for RuBisCOfrom the photosynthetic bacterium Rhodospirillum rubrum. Thus, the photosynthetic RuBisCO from R. rubrum retains the ability to function in the methionine salvage pathway in B. subtilis.

Amino Acid Sequence↗
PubMed2003

Direct observation of guanine radical cation deprotonation in duplex DNA using pulse radiolysis.

The dynamics of one-electron oxidation of guanine (G) base mononucleotide and that in DNA have been investigated by pulse radiolysis. The radical cation (G+*) of deoxyguanosine (dG), produced by oxidation with SO(4)-*, rapidly deprotonates to form the neutral G radical (G(-H)*) with a rate constant of 1.8 x 10(7) s(-1) at pH 7.0, as judged from transient spectroscopy. With experiments using different double-stranded oligonucleotides containing G, GG, and GGG sequences, the absorbance increases at 625 nm, characteristic of formation of the G(-H)*, were found to consist of two phases. The rate constants of the faster ( approximately 1.3 x 10(7) s(-1)) and slower phases ( approximately 3.0 x 10(6) s(-1)) were similar for the different oligonucleotides. On the other hand, in the oligonucleotide containing G located at the 5'- and 3'-terminal positions, only the faster phase was seen. These results suggest that the lifetime of the radical cation of the G:C base pair (GC+*), depending on its location in the DNA chain, is longer than that of free dG. In addition, the absorption spectral intermediates showed that hole transport to a specific G site within a 12-13mer double-stranded oligonucleotide is complete within 50 ns; that is, the rate of hole transport over 20 A is >10(7) s(-1).

Cations↗
PubMed2003

An RNA-modifying enzyme that governs both the codon and amino acid specificities of isoleucine tRNA.

The AUA codon-specific isoleucine tRNA (tRNA(Ile)) in eubacteria has the posttranscriptionally modified nucleoside lysidine (L) at the wobble position of the anticodon (position 34). This modification is a lysine-containing cytidine derivative that converts both the codon specificity of tRNA(Ile) from AUG to AUA and its amino acid specificity from methionine to isoleucine. We identified an essential gene (tilS; tRNA(Ile)-lysidine synthetase) that is responsible for lysidine formation in both Bacillus subtilis and Escherichia coli. The recombinant enzyme complexed specifically with tRNA(Ile) and synthesized L by utilizing ATP and lysine as substrates. The lysidine synthesis of this enzyme was shown to directly convert the amino acid specificity of tRNA(Ile) from methionine to isoleucine in vitro. Partial inactivation of tilS in vivo resulted in an AUA codon-dependent translational defect, which supports the notion that TilS is an RNA-modifying enzyme that plays a critical role in the accurate decoding of genetic information.

Amino Acid Sequence↗
PubMed2003

The BceRS two-component regulatory system induces expression of the bacitracin transporter, BceAB, in Bacillus subtilis.

BceA and bceB encode a nucleotide-binding domain (NBD) and membrane-spanning domain (MSD) subunit, respectively, of an ATP-binding cassette (ABC) transporter in Bacillus subtilis. Disruption of these genes resulted in hypersensitivity to bacitracin, a peptide antibiotic that is non-ribosomally synthesized in some strains of Bacillus. Northern hybridization analyses showed that expression of the bceAB operon is induced by bacitracin present in the growth medium. The bceRS genes encoding a two-component regulatory system are located immediately upstream of bceAB. Deletion analyses of the bceAB promoter together with DNase I footprinting experiments revealed that a sensor kinase, BceS, responds to extracellular bacitracin either directly or indirectly and transmits a signal to a cognate response regulator, BceR. The regulator binds directly to the upstream region of the bceAB promoter and upregulates the expression of bceAB genes. The bcrC gene product is additionally involved in bacitracin resistance. The expression of bcrC is dependent on the ECF sigma factors, sigmaM and sigmaX, but not on the BceRS two-component system. In view of these results, possible roles of BceA, BceB and BcrC in bacitracin resistance of B. subtilis 168 are discussed.

ATP-Binding Cassette Transporters↗
PubMed2003

Chryseobacterium miricola sp. nov., a novel species isolated from condensation water of space station Mir.

Classification of strain W3-B1, which was isolated from condensation water in the Russian space laboratory Mir, was investigated by a polyphasic taxonomic approach. Cells of strain W3-B1 were nonmotile, asporogenous, gram-negative slender rods with rounded ends. 16S rRNA gene sequence analysis indicated that organism should be placed in the genus Chryseobacterium. This organism contains menaquinone MK-6 as the predominent isoprenoid quinone and 3-OH iso 17:0 (40%), iso 15:0 (33%) as the major fatty acids. Phylogenetically, the nearest relative of strain W3-B1 is Chryseobacterium meningosepticum with sequence similarity of 98.4%, but DNA-DNA hybridization resulted in similarity values of only 52.3%. The G+C mol% is 34.6 mol%. Based upon results obtained by morphological, biochemical, chemotaxonomic, and molecular methods, strain W3-B1 was clearly distinguishable from other Chryseobacterium species. For these reasons, a novel species of family Flavobacteriaceae is proposed; strain W3-B1(T) (= GTC 862(T) = JCM 11413(T) = DSM 14571(T)) is the type strain.

Base Composition↗
PubMed2003

Bacterial ceramides and sphingophospholipids induce apoptosis of human leukaemic cells.

The genus Sphingobacterium, whose members are Gram-negative non-fermentative rods, possesses ceramides and related sphingophospholipids (SPLs) with isoheptadecasphinganine and 2-hydroxy or non-hydroxy isopentadecanoic acid. This paper reports evidence that ceramides isolated from Sphingobacterium spiritivorum ATCC 33861 induce endonucleolytic DNA cleavage in human myeloid leukaemia HL-60 cells in vitro, which is the primary characteristic biochemical marker for apoptosis or programmed cell death. Ceramides and SPLs also induced DNA fragmentation and caspase-3 activation, followed by changes in morphology, such as alterations in the size of nuclei and cells, and cell cycle shortening. Apoptotic activity correlated with the ceramide structure. Ceramide with a 2-hydroxy fatty acid showed stronger apoptotic activity than ceramide with a non-hydroxy fatty acid. Furthermore, the major five SPLs (ceramide phosphorylethanolamine-1 and -2, ceramide phosphorylinositol-1 and -2, and ceramide phosphorylmannose-1) showed apoptosis-inducing activity in HL-60 cells, indicating that the ceramide moiety of the SPLs plays a crucial role as the intracellular second messenger but that their hydrophilicity is less important in this regard. The hydrophilic part of SPLs may play a role in other cellular response systems. The involvement of Fas antigen was implicated in the apoptotic event since Fas antigen expression was observed after 3 or 4 h stimulation of HL-60 cells with bacterial ceramides. However, a time-course study for caspase-3 activation indicated maximal activity at 1 h after stimulation with bacterial ceramides, suggesting that two (or possibly more) mechanisms of signal transduction, Fas-dependent and Fas-independent, may be involved. Fas antigen expression and caspase-3 activation by five kinds of SPLs were observed after 3 or 4 h. These results indicate that there is a difference in the response of HL-60 cells to bacterial ceramides and SPLs.

Apoptosis↗
PubMed2003

The Bacillus subtilis YufLM two-component system regulates the expression of the malate transporters MaeN (YufR) and YflS, and is essential for utilization of malate in minimal medium.

The Gram-positive bacterium Bacillus subtilis has a complete set of enzymes for the tricarboxylic acid (TCA) cycle and can grow aerobically using most of the TCA cycle intermediates (malate, fumarate, succinate and citrate) as a sole carbon source. The B. subtilis genome sequence contains three paralogous two-component regulatory systems, CitST, DctSR and YufLM. CitST and DctSR activate the expression of a transporter of the Mg(2+)-citrate complex (CitM) and a fumarate and succinate transporter (DctP), respectively. These findings prompted an investigation of whether the YufL sensor and its cognate regulator, YufM, play a role in malate uptake. This paper reports that the YufM regulator shows in vitro binding to the promoter region of two malate transporter genes, maeN and yflS, and is responsible for inducing their expression in vivo. It was also found that inactivation of the yufM or maeN genes resulted in bacteria that could not grow in a minimal salts medium containing malate as a sole carbon source, indicating that the induction of the MaeN transporter by the YufM regulator is essential for the utilization of malate as a carbon source. Inactivation of the yufL gene resulted in the constitutive expression of MaeN. This expression was suppressed by reintroduction of the kinase domain of YufL, indicating that the YufL sensor is required for proper signal detection and signalling specificity. The authors propose that a phosphatase activity of YufL plays an important role in the YufLM two-component regulatory system. The studies reported here have revealed that members of a set of paralogous two-component regulatory systems in B. subtilis, CitST, DctSR and YufLM, are involved in a related function--uptake (and metabolism) of the TCA cycle intermediates--but with distinct substrate specificities.

Bacillus subtilis↗
PubMed2003

Bacillus subtilis spoVIF (yjcC) gene, involved in coat assembly and spore resistance.

In systematic screening four sporulation-specific genes, yjcA, yjcB, yjcZ and yjcC, of unknown function were found in Bacillus subtilis. These genes are located just upstream of the cotVWXYZ gene cluster oriented in the opposite direction. Northern blot analysis showed that yjcA was transcribed by the SigE RNA polymerase beginning 2 h (t(2)) after the onset of sporulation, and yjcB, yjcZ and yjcC were transcribed by the SigK RNA polymerase beginning at t(4) of sporulation. The transcription of yjcZ was dependent on SigK and GerE. The consensus sequences of the appropriate sigma factors were found upstream of each gene. There were putative GerE-binding sites upstream of yjcZ. Insertional inactivation of the yjcC gene resulted in a reduction in resistance of the mutant spores to lysozyme and heat. Transmission electron microscopic examination of yjcC spores revealed a defect of sporulation at stage VI, resulting in loss of spore coats. These results suggest that YjcC is involved in assembly of spore coat proteins that have roles in lysozyme resistance. It is proposed that yjcC should be renamed as spoVIF.

Amino Acid Sequence↗
PubMed2003

Bacillus subtilis diacylglycerol kinase (DgkA) enhances efficient sporulation.

The sn-1,2-diacylglycerol kinase homologue gene, dgkA, is a sporulation gene indispensable for the maintenance of spore stability and viability in Bacillus subtilis. After 6 h of growth in resuspension medium, the endospore morphology of the dgkA mutant by standard phase-contrast microscopy was normal; however, after 9 h, the endospores appeared mostly dark by phase-contrast microscopy, suggesting a defect in the spores. Moreover, electron microscopic studies revealed an abnormal cortex structure in mutant endospores 6 h after the onset of sporulation, an indication of cortex degeneration. In addition, a significant decrease in the dipicolinic acid content of mutant spores was observed. We also found that dgkA is expressed mainly during the vegetative phase. It seems likely that either the DgkA produced during growth prepares the cell for an essential step in sporulation or the enzyme persists into sporulation and performs an essential function.

Bacillus subtilis↗
PubMed2003

Effect of lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin on the susceptibility to Listeria infection.

We studied the effect of lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the susceptibility to Listeria infection of offspring in C57BL /6NCji mice. The offspring were nursed by TCDD-treated dams and exposed to TCDD from birth to weaning via milk. The exposure had little effect on the weights of immune organs and the spleen or the thymus cell population in the dams and offspring, but it enhanced the production of tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) in the serum after Listeria infection. The clearance of Listeria monocytogenes from the spleen was impaired in the off-spring. These results suggest that the exposure to TCDD of the offspring via milk disrupted the host resistance of the offspring, even though the main immune parameters were unchanged.

Animals↗
PubMed2003

Inferring gene regulatory networks from time-ordered gene expression data of Bacillus subtilis using differential equations.

We describe a new method to infer a gene regulatory network, in terms of a linear system of differential equations, from time course gene expression data. As biologically the gene regulatory network is known to be sparse, we expect most coefficients in such a linear system of differential equations to be zero. In previously proposed methods, the number of nonzero coefficients in the system was limited based on ad hoc assumptions. Instead, we propose to infer the degree of sparseness of the gene regulatory network from the data, where we use Akaike's Information Criterion to determine which coefficients are nonzero. We apply our method to MMGE time course data of Bacillus subtilis.

Bacillus subtilis↗
PubMed2003

[Up-to-date understanding of tuberculosis immunity].

This symposium was organized to provide the up-to-date knowledge on tuberculosis immunity, especially on the understanding of cytokines or Th1 cells involved in pathophysiology/protective immunity and vaccine development. Dr. Kazuo Kobayashi (Osaka City Univ.) reported their findings on the immune response to bioactive lipid component from M. tuberculosis, trehalose-dimycolate (TDM) and sulfolipid (SL) in mice. Their unique and novel finding was that TDM is capable of inducing T-dependent immune response in euthymic mice. The specific immune response in TDM-immune mice was consisting of CD4+ cell response and expression of chemokines, inflammatory cytokines and then TH1-related cytokines. In contrast, SL did not show such an activity. TDM may be one of the protective antigens and may modulate the specific immune response of the host. Dr. Isamu Sugawara's group (JATA) has examined the involvement of various cytokines in the host response to aerosolic infection with virulent strain of M. tuberculosis by using cytokine-knockout mice. The single deletion of IFN-gamma or TNF alpha resulted in a severe lesion of multiple necrosis without granuloma, and cytokine mRNA level other than knocked out cytokine was normal, suggesting that IFN-gamma and TNF alpha are principally important cytokines. In knockout mice for IL-12 or IL-18, necrotic lesion was not induced after infection and the pathological change was not so significant as in IFN-gamma/TNF alpha knockout mice. By using IFN-gamma knockout mice, it became possible to generate a granulomatous lesion with central necrosis and cavity resembling the lesion in humans. These mouse model appeared to be useful in the analysis of pathophysiology of human tuberculosis. Dr. Kazuyoshi Kawakami (Ryukyu Univ.) reported the importance of TH1 cytokines in anti-tuberculous immunity. By using IL-12, IL-18 knockout mice or double knockout mice, it was shown that IL-12 exhibits more important role than IL-18 in the protection. A possible contribution of IL-23 was also suggested. In most of the clinical cases of tuberculosis, the production of IL-12, IL-18 and IFN-gamma is increased, however, the group of relatively lower cytokine production did not respond well to the treatment. In addition, the plasma level of one of the chemokines, IP-10, was shown to be an indicator for the severity of the disease. Thus, some cytokines appear to be employable for the novel treatment in the near future. Dr. Saburo Yamamoto (NIH) summarized the recent advance in the understanding of biological function of CpG motifs. Immunostimulatory DNA is effective in the modulation of TH1/TH2 polarity and the enhancement of protective immunity to M. tuberulosis in animals. CpG motif (immunostimulatory DNA) appears to exert its activity by signaling cascade via TLR9 resulting in NF-kappa B activation and cytokine gene expression. Analysis of basic mechanism of action by CpG motif should pave the way to the clinical application in the future. Dr. Masaji Okada (Kinki Chuo Hospital) reported the current situation in the development of novel vaccines against tuberculosis. They have extensively constructed and examined the efficacy of various types of vaccines including subunit, DNA and recombinant BCG vaccines. Various vector systems have been tested for DNA vaccine. As immunizing antigens, a-Ag, ESAT-6, HSP65, 38kD-lipoprotein and so on have been employed. A large body of experimental data are accumulating for final evaluation, and among them, it is noteworthy to mention that HSP65DNA + IL-12DNA was 100 times more effective than conventional BCG in animal model. Among subunit vaccines, Mtb72f vaccine appears to be one of the promising candidates. In addition to the trial with various candidates, they have established a new mouse model, SCID/human PBL. This model animal has been employed for the development of vaccine effective for the induction of ESAT-6-specific human T cells.

Animals↗
PubMed2002

Serodiagnosis of pulmonary disease due to Mycobacterium avium complex with an enzyme immunoassay that uses a mixture of glycopeptidolipid antigens.

It is difficult to distinguish pulmonary disease due to Mycobacterium avium complex (MAC) from that due to other mycobacteria, such as Mycobacterium tuberculosis and Mycobacterium kansasii. We developed an enzyme immunoassay (EIA) for diagnosis of MAC pulmonary diseases that uses glycopeptidolipid (GPL) antigens specific for MAC, and we used it for diagnosis in immunocompetent patients. The mean optical densities (+/- standard deviation) of serum immunoglobulin G antibodies to GPLs in patients with MAC disease, MAC colonization, M. kansasii disease, and tuberculosis and in healthy subjects were 0.778+/-0.784, 0.042+/-0.035, 0.059+/-0.035, 0.071+/-0.035, and 0.030+/-0.027, respectively. A significant increase in the level of anti-GPL antibodies was detected in patients with MAC disease. The level of anti-GPL antibodies reflected disease activity, because the level was decreased in culture-negative patients who had conversion of culture results. When a cutoff level of seropositivity (0.119) was defined, the sensitivity of EIA for diagnosis of MAC disease was 92.3%, and the specificity was 96.7%. Measurement of serum anti-GPL antibodies is useful for both the diagnosis of and assessment of activity in MAC disease.

Adolescent↗
PubMed2002

Spectroscopic and functional characterization of Cu-containing nitrite reductase from Hyphomicrobium denitrificans A3151.

The Cu-containing nitrite reductase from Hyphomicrobium denitrificans (HydNIR) has been spectroscopically and functionally characterized. The visible absorption spectrum implies that the enzyme has two type 1 Cu ions in one subunit (ca. 50 kDa). The electron paramagnetic resonance (EPR) spectrum of HydNIR is simulated assuming the sum of three distinct S = 1/2 systems: two type 1 Cu signals (axial and rhombic symmetries) and one type 2 Cu signal. The intramolecular electron transfer reaction from the type 1 Cu to the type 2 Cu at pH 6.0 does not occur in the absence of nitrite, but a very slow electron transfer reaction is observed in the presence of nitrite. The apparent first-order rate constants for the intramolecular electron transfer reactions (k(ET(intra))) in the presence of nitrite and also the apparent catalytic rate constants (k(cat)) of HydNIR decrease gradually with increasing pH in the range of pH 4.5-7.5. These pH profiles are substantially similar to each other, suggesting that the intramolecular electron transfer process is linked to the subsequent nitrite reduction process.

Bacterial Proteins↗
PubMed2002

Enzymatic reaction of hydrogen peroxide-dependent peroxygenase cytochrome P450s: kinetic deuterium isotope effects and analyses by resonance Raman spectroscopy.

Cytochromes P450SP(alpha) (CYP152B1) and P450BS(beta) (CYP152A1), which are isolated from Sphingomonas paucimobilis and Bacillus subtilis, respectively, belong to the P450 superfamily, but catalyze hydroxylation reactions, in which an oxygen atom from H2O2 is efficiently introduced into fatty acids (e.g., myristic acid). P450SP(alpha) produces the alpha-hydroxylated (alpha-OH) products at 100%, while P450BS(beta) produces alpha- and beta-hydroxylated (beta-OH) products at 33 and 67%, respectively. Using deuterium-substituted fatty acids ([2,2-d2]-myristic acid and d27-myristic acid) as a substrate, the peroxygenase reactions of the two bacterial P450s were investigated. In the P450SP(alpha) reaction, we observed an intermolecular noncompetitive kinetic isotope effect on Vmax (DV = 4.1) when [2,2-d2]-myristic acid was used, suggesting that an isotopically sensitive step involving the alpha-hydrogen of the fatty acid is present in the catalytic cycle. On the other hand, D(V/K) was masked, in sharp contrast to the features of usual monooxygenases P450. The characteristic kinetic features can be interpreted in terms of the faster product formation than the substrate dissociation. A similar kinetic isotope effect was observed [DV = 4.9, D(V/K) approximately 1] for the P450BS(beta) reaction, when d27-myristic acid was used as a substrate, indicating that the reaction mechanism is the same for both peroxygenases. The resonance Raman spectral data of P450BS(beta) in the ferric and ferrous-CO forms in the presence and absence of myristic acid demonstrated that the catalytic pocket of the enzyme is polar, so that the location of the carboxylate of the substrate close to the sixth ligand of the heme could be allowed. On the basis of these results on the kinetic isotope effects and spectroscopy, we discuss the possible mechanisms of the alpha- and beta-hydroxylation of fatty acids catalyzed by peroxygenases P450SP(alpha) and P450BS(beta).

Bacillus subtilis↗
PubMed2002

Interaction of monocytes with vascular endothelial cells synergistically induces interferon gamma-inducible protein 10 expression through activation of specific cell surface molecules and cytokines.

To further understand the regulatory mechanisms involved in the process of angiogenesis, the present study was designed to determine the expression and regulation of interferon gamma-inducible protein 10 (IP-10) in peripheral blood monocytes and human umbilical vein endothelial cells (HUVECs). We found that the interaction of monocytes with HUVECs resulted in synergistic increases in IP-10 expression and secretion, which consequently inhibited endothelial tube formation in vitro. Induction of IP-10 was mediated via specific cell surface molecules, as indicated by the finding that IP-10 secretion was significantly inhibited by anti-CD40 ligand antibody, and to a lesser extent by anti-CD40 antibody. Furthermore, we examined the effects of soluble mediators, such as inflammatory and immune cytokines on IP-10 secretion. Addition of interleukin (IL)-1, as well as interferon gamma, induced a marked augmentation of IP-10 secretion by unstimulated monocytes, unstimulated HUVECs, and co-cultures of the two cell types. In contrast, IL-10, recognized as an anti-inflammatory cytokine, significantly inhibited IP-10 secretion by co-cultures. Our results suggest that the interaction of monocytes with endothelial cells results in synergistic increases in IP-10 expression and secretion, which contribute to the regulation of angiogenesis and initiation of inflammatory vascular diseases.

CD40 Antigens↗