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N Mutoh

Publications and source records attributed to N Mutoh.

49 records · Page 3Linked to original sources
PubMed1986

Acquisition of maltose chemotaxis in Salmonella typhimurium by the introduction of the Escherichia coli chemosensory transducer gene.

Escherichia coli and Salmonella typhimurium are closely related species. However, E. coli cells show maltose chemotaxis but S. typhimurium cells do not. When an E. coli chemotransducer gene (tarE), the product of which is required for both aspartate and maltose chemotaxis, was introduced by using a plasmid vector into S. typhimurium cells with a defect in the corresponding gene (tarS), the transformant cells acquired the ability for both aspartate and maltose chemotaxis. In contrast, when the tars gene was introduced into tarE-deficient E. coli cells, the transformant cells acquired aspartate chemotaxis but not maltose chemotaxis. These results indicate that the absense of maltose chemotaxis in S. typhimurium is a consequence of the properties of the tars gene product.

Aspartic Acid↗
PubMed1986

Characterization of Escherichia coli chemotaxis receptor mutants with null phenotypes.

Hydroxylamine mutagenesis was used to alter the tar gene that encodes the transmembrane Tar protein required for chemotaxis. Mutants defective in chemotaxis were selected, and the mutation was characterized by DNA sequencing. Two classes of mutations were found: nonsense and missense. The nonsense mutations were distributed throughout the gene, while the missense mutations were found to cluster in a region that includes 185 amino acids at the C-terminal end of the Tar protein. Partial characterization of mutant phenotypes suggested that some are completely defective in signaling while responding to attractants and repellents by differential methylation. Other mutants are undermethylated and constantly tumble, while yet another class of mutants is overmethylated and biased toward constant swimming with little or no tumbling. These mutants will be useful in experiments designed to understand the mechanism of chemotaxis.

Amino Acids↗
PubMed1983

Sensory transducers of E. coli are composed of discrete structural and functional domains.

The tar and tsr genes of E. coli encode functionally analogous transducer proteins that mediate two distinct classes of chemotactic response. The tap gene lies adjacent to tar, and is thought to encode another transducer protein. We present here the complete nucleotide sequence of the tar-tap region of the E. coli genome, together with a comparative analysis of the sequences of the Tar, Tap, and Tsr proteins. The proteins appear to have a simple transmembrane structure consisting of an extracytoplasmic amino-terminal domain, a membrane-spanning domain, and an intracellular carboxy-terminal domain. The carboxy-terminal domains of three proteins possess highly homologous sequences and contain sites of methylation involved in sensory adaptation, while the amino-terminal sequences are only distantly related to one another, consistent with their serving as chemoreceptor domains that have diverged functionally.

Amino Acid Sequence↗
PubMed1983

Mutation causing overproduction of outer membrane protein OmpF and suppression of OmpC synthesis in Escherichia coli.

A novel mutation affecting the synthesis of major outer membrane proteins OmpF and OmpC in Escherichia coli K-12 is described. The mutation resulted in overproduction of the OmpF protein with concomitant suppression of OmpC synthesis. This mutation, designated as ompFp100, was mapped at 21 min on the E. coli chromosome map with the gene order aroA-aspC-ompF4-ompFp100-asnS-pyrD. This mutation was cis-dominant to the expression of the ompF gene. In addition, the direction of the mRNA transcription of the ompF gene was from asnS to aspC. These results strongly indicate that ompFp100 is a promoter mutation of the ompF gene. Introduction of an ompF mutation, which causes the disappearance of the OmpF protein, into strains carrying the ompFp100 mutation resulted in the reappearance of the OmpC protein in the outer membrane. Addition of a high concentration of sucrose to the medium, which suppresses the OmpF synthesis and stimulates the OmpC synthesis in the wild-type strain, resulted in the reappearance of the OmpC protein in the ompFp100 mutant with concomitant suppression of the overproduction of the OmpF protein. These results suggest that suppression of OmpC synthesis in the ompFp100 mutant is due to overproduction of the OmpF protein.

Bacterial Outer Membrane Proteins↗
PubMed1982

Primary structure of the ompF gene that codes for a major outer membrane protein of Escherichia coli K-12.

The nucleotide sequence of the ompF gene coding for a major outer membrane protein of Escherichia coli K-12 has been determined and the amino acid sequence of the OmpF protein was deduced from it. The OmpF protein contains 340 amino acid residues, and is produced from a precursor having 22 extra amino acid residues, the signal peptide, at the amino terminus. The expected secondary structure of the OmpF protein had a high beta-sheet content with a low alpha-helix content. The promoter region and the transcription termination region of the ompF gene had a significantly high AT content, while the AT content of the coding region was about the same as the average AT content of the E. coli chromosome. Following the termination codon, a typical rho-independent transcription termination signal was observed. The codon usage in the ompF gene was highly nonrandom; the codons preferably utilized are those recognized by the most abundant species of isoaccepting tRNAs or those, among synonymous codons recognized by the same tRNA, that can interact more properly with the anticodon.

Amino Acid Sequence↗
PubMed1981

Use of lipophilic cation-permeable mutants for measurement of transmembrane electrical potential in metabolizing cells of Escherichia coli.

Some lipopolysaccharide-defective mutants of Escherichia coli showed, without ethylenediaminetetraacetic acid treatment, a quick and high uptake of lipophilic cations such as triphenylmethylphosphonium and tetraphenylphosphonium. The rate and amount of uptake were comparable to those of an ethylenediaminetetraacetic acid-treated wild type. Transmembrane electrical potential, which was calculated from the distribution of these lipophilic cations between the inside and outside of the mutant cells, was about -150 mV at pH 7.5 and showed a strong dependency on the external pH. One of the E. coli mutants, the acrA mutant, was found to be also permeable to dicyclohexylcarbodiimide, an H+-adenosine triphosphatase inhibitor, and 1-anilino-8-naphthalene sulfonate, a fluorescent dye. The acrA mutant was vigorously motile and highly sensitive to many bacteriophages and colicins. Thus, the acrA mutant is quite useful for the quantitative measurement of transmembrane electrical potential by lipophilic cations in intact and metabolizing cells especially in relation to motility and actions of colicins and bacteriophages.

Cell Membrane Permeability↗
PubMed1978

Role of lipopolysaccharide and outer membrane protein of Escherichia coli K-12 in the receptor activity for bacteriophage T4.

Lipopolysaccharide isolated from Escherichia coli K-12 did not inactivate phage T4, although the cell envelopes with 1% sodium deoxycholate resulted in the release of cytoplasmic membrane proteins, 70% of the lipopolysaccharide, and almost all of the phospholipid. The reconstitution of phage receptor activity was achieved from deoxycholate-soluble and -insoluble fractions by dialysis against a solution of magnesium chloride. Lipopolysaccharide was the only essential component in the deoxycholate-soluble fraction. PhageT4-resistant mutants YA21-6 and YA21-82, having defects in the deoxycholate-soluble and -insoluble fractions, respectively, were isolated. The deoxycholate-soluble fraction of YA21-6 possessed heptoseless lipopolysaccharide, and this defect was responsible for the phage resistance. The deoxycholate-insoluble fraction of YA21-82 lacked outer membrane protein O-8. The addition of O-8 to this fraction together with the wild-type lipopolysaccharide resulted in the appearance of the receptor activity. Furthermore, the reconstitution was successfully achieved with only O-8 and the wild-type lipopolysaccharide, indicating that O-8 was an essential component in the deoxycholate-insoluble fraction.

Bacterial Proteins↗
PubMedDate not supplied

Two pathways in the biosynthesis of cadystins (gamma EC)nG in the cell-free system of the fission yeast.

Small metal-binding peptides, cadystins, with the general structure of (gamma-Glu-Cys)n-Gly ((gamma EC)nG), were synthesized in a cell-free system of fission yeast to examine the in vivo synthetic pathway. The crude enzyme for cadystin synthesis was prepared by ammonium sulfate precipitation (75% saturation) from the 120,000 x g supernatant of the cell extract, and the excess salt in the enzyme fraction was removed by Sephadex gel filtration. Using this crude enzyme fraction, it was shown that there were two pathways for cadystin biosynthesis. One pathway is gamma-Glu-Cys (gamma EC) dipeptidyl transfer from both glutathione (gamma ECG) and cadystins to glutathione and cadystins. The other one is gamma EC polymerization from (gamma EC)n and glutathione to (gamma EC)n + i, followed by glycine addition with glutathione synthetase.

Amino Acid Sequence↗
PubMedDate not supplied

Glutathione synthetase from the fission yeast. Purification and its unique heteromeric subunit structure.

Glutathione (GSH) synthetase (EC 6.3.2.3) was purified from the fission yeast Schizosaccharomyces pombe L972h- and from the GSH synthetase deficient mutant MN101/pYS41, which harbors a plasmid containing the GSH synthetase gene of the fission yeast. GSH synthetase is expressed at 10 times higher the amount in MN101/pYS41 than in wild-type L972h-. The purified enzyme gave a single band on polyacrylamide gel electrophoresis in the absence of sodium dodecyl sulfate (native PAGE). The molecular weight of this enzyme was determined to be 1.2 x 10(5) by Sepharose CL-6B gel filtration. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE) revealed that this enzyme was composed of two kinds of subunits, A (M(r) = 33 x 10(3)) and B (M(r) = 26 x 10(3)), and existed as a heterotetramer (A2B2). The enzyme purified from the wild-type fission yeast, which did not harbor the plasmid, showed the same electrophoretic mobilities on both native PAGE and SDS-PAGE and similar catalytic properties under standard conditions. This enzyme is most active at 45 degrees C and pH 8.0-8.5 with 20 mM Mg2+ + 10 mM ATP and 50 mM K+. The strict requirement for the monovalent cation is rather specific for the enzymes from yeasts. The presence of sugar components in the enzyme is also observed, similar to that in the rat kidney enzyme.

Animals↗