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Biomedical subjects

K Yoda

Publications and source records attributed to K Yoda.

At least 109 records · Page 6Linked to original sources
PubMed1987

Molecular cloning and nucleotide sequence of the 3-isopropylmalate dehydrogenase gene of Candida utilis.

A 3-isopropylmalate dehydrogenase (3-IMDH, EC 1.1.1.85) gene was cloned from a gene library of Candida utilis. One of the plasmids, pYKL30, could complement Escherichia coli leuB and Saccharomyces cerevisiae leu2 auxotrophs; a 2.2 kb HindIII fragment subcloned in pBR322 could still complement the leuB mutation. Southern hybridization confirmed that this fragment was derived from C. utilis. An open reading frame of 1089 bp that corresponded to a polypeptide of 363 amino acids, one residue shorter than the 3-IMDH of S. cerevisiae, was found in the cloned fragment. The homology between the 3-IMDHs of C. utilis and S. cerevisiae was 76.2% in nucleotides and 85.4% in amino acids. In contrast, the homology between the 3-IMDHs of C. utilis and Thermus thermophilus was much smaller and was restricted to some regions of the gene.

3-Isopropylmalate Dehydrogenase↗
PubMed1987

Secretion of Bacillus subtilis alpha-amylase in the periplasmic space of Escherichia coli.

The Bacillus subtilis alpha-amylase structural gene (amyE) lacking its own signal peptide coding sequence was joined to the end of the Escherichia coli alkaline phosphatase (phoA) signal peptide coding sequence by using the technique of oligonucleotide-directed site-specific deletion. On induction of the phoA promoter, the B. subtilis alpha-amylase was expressed and almost all the activity was found in the periplasmic space of E. coli. The sequence of the five amino-terminal amino acids of the secreted polypeptide was Glu-Thr-Ala-Asn-Lys-, and thus the fused protein was correctly processed by the E. coli signal peptidase at the end of the phoA signal peptide.

Bacillus subtilis↗
PubMed1986

Cooperative inhibitory effect of follicular fluid and cAMP on hamster oocyte maturation.

Porcine or human follicular fluid inhibited the spontaneous maturation of isolated hamster oocytes in vitro during the first 1.5 h of culture. Moreover, the presence of 50% follicular fluid combined with 100 microM dbcAMP cooperatively reduced the incidence of germinal vesicle breakdown. The addition of FSH also inhibited the resumption of meiosis, and the presence of LH did not overcome the inhibitory effects of follicular fluid and tended to impede isolated hamster oocyte maturation in vitro.

Animals↗
PubMed1986

Amplification of the amyE-tmrB region on the chromosome in tunicamycin-resistant cells of Bacillus subtilis.

In a class of tunicamycin-resistant mutants (tmrA7) of Bacillus subtilis, the production of extracellular alpha-amylase is increased by about five fold. The tmrA7 characteristics (tunicamycin resistance and hyperproduction of extracellular alpha-amylase) can be transferred to recipient cells by transformation. In the transformants and the original tmrA7 mutant, typical amplification of the region from 4 kb upstream of the amyE gene to the tmrB gene on the chromosome was detected. The repeating unit, 16 kb in size, repeats tandemly about five and ten times in the mutant and transformants, respectively, and the alpha-amylase production is proportional to the copy number of the amyE gene. Simultaneous amplification of the tmrB gene, which is responsible for tunicamycin resistance in the multicopy state, and the alpha-amylase structural gene (amyE) seems to be the cause of the pleiotropy of the tmrA7 mutation.

Bacillus subtilis↗
PubMed1986

Amplification and deletion of the amyE+-tmrB+ gene region in a Bacillus subtilis recombinant-phage genome by the tmrA7 mutation.

A 22.4-kilobase DNA fragment containing the tmrA7-amyR2-amyE+-tmrB+-aroI+ region of the Bacillus subtilis N7 chromosomal DNA was cloned into a recombinant B. subtilis bacteriophage, p11-AA248. The amyE+-tmrB+ gene region, approximately 12.6 kilobases, in the phage genome was amplified in a tunicamycin-resistant (Tmr) Amy+ AroI+ transductant of B. subtilis by p11-AA248. On the other hand, the amyE+-tmrB+ region in the genomes of 80 to 90% of the phage particles was deleted when the phages were induced from the Tmr Amy+ AroI+ transductants by treatment with 1.0 micrograms of mitomycin C per ml. From analyses of the physical maps and DNA nucleotide sequences in the junction region of the deleted phage genome and the parental DNA fragments, it is suggested that the deletion occurred within a direct repeat sequence composed of 18 base pairs. The endpoints of the amplified gene region seemed to be closely related to both terminal regions of the deleted DNA.

Bacillus subtilis↗
PubMed1986

Essential structure in the cloned transforming DNA that induces gene amplification of the Bacillus subtilis amyE-tmrB region.

Bacillus subtilis B7, a mutant which acquired gene amplification of the amyE-tmrB region, showed, as a result, hyperproductivity (about a 5- to 10-fold increase) of alpha-amylase and tunicamycin resistance. The mutational character was transferred to recipient cells by competence transformation. A 14-kilobase (kb) EcoRI chromosomal DNA fragment of strain B7 was found to have the transforming activity. We cloned a 6.4-kb EcoRI fragment on a phage vector lambda Charon 4A through a spontaneous deletion of 7.6 kb from the 14-kb fragment and subcloned a 1.6-kb HindIII fragment on pGR71. The cloned 6.4-kb EcoRI and 1.6-kb HindIII fragments retained the transforming activity of inducing gene amplification of the amyE-tmrB region. At the junction point (J) of the repeating units (16 kb), the tmrB gene was linked to a DNA region (M) located 4 kb upstream of amyE. The essential structure of the cloned, transforming (gene amplification-inducing) DNA was deduced to be that around J. The subcloned 1.6-kb HindIII fragment that retained the transforming activity was shown to be almost solely composed of the tmrB-J-M region. In addition, the DNA sequence around J was determined.

Bacillus subtilis↗
PubMed1985

Evidence that discontinuous DNA replication in Escherichia coli is primed by approximately 10 to 12 residues of RNA starting with a purine.

Intact primer RNA for discontinuous DNA replication of Escherichia coli has been detected by specific labeling in vitro of its 5'-terminal tri- (or di-) phosphate group with vaccinia guanylyltransferase and [alpha-32P]GTP. A mutant defective either in RNase H or in both RNase H and DNA polymerase I accumulated about 10 or 30 times more intact primer RNA, respectively, than wild-type cells. The primers started with purine in an A to G ratio of 5 and the most abundant 5'-terminal dinucleotide sequence was (p)ppA-Pu. The chain length of the intact primer RNA was approximately 10 to 12 nucleotide residues. The structural properties of the E. coli primer RNa resemble those of the eukaryotic primer RNA.

Base Sequence↗
PubMed1985

Structure of a processed gene of mouse cytoplasmic gamma-actin transposed into a BAM5 sequence: insertion has created 13 base-pair direct repeats.

We have cloned a processed gene related to cytoplasmic gamma-actin from a phage library of mouse brain DNA. The gene lacks introns, carries a poly(dA) tract of 19-base pairs at the 3' end and is flanked by 13-base-pair direct repeats. Southern blotting experiments indicated that this segment is embedded in highly repetitive and strongly methylated sequences. Direct comparison of the flanking sequences with those of mouse repetitive DNAs revealed that this processed gene is integrated into a BAM5 family, which is one of the repetitive sequences of the mouse genome. In addition, it was shown that the direct repeats were created by a duplication of the corresponding 13-bp stretch of the BAM5 DNA as a consequence of the integration of this segment. These results strongly suggest that the processed gamma-actin gene has been conveyed to this locus through a poly(A)+ RNA intermediate by a transposon-like mechanism.

Actins↗
PubMed1985

Synthesis and secretion of human epidermal growth factor by Escherichia coli.

A synthetic gene for human epidermal growth factor (hEGF) was joined to a sequence encoding the signal peptide of Escherichia coli alkaline phosphatase. This hybrid gene was placed under the control of the alkaline phosphatase gene (phoA) promoter in a recombinant plasmid, which was used to transfect E. coli. The hybrid protein that was expressed in host cells under conditions of phosphate limitation was processed accurately during the secretion process, and mature hEGF was recovered in the periplasmic fraction. On the other hand, no EGF was detected in the periplasmic space when the synthetic hEGF gene was not accompanied by the phoA signal sequence.

Alkaline Phosphatase↗
PubMed1985

Secretion of human interferon-alpha induced by using secretion vectors containing a promoter and signal sequence of alkaline phosphatase gene of Escherichia coli.

We constructed a new vector containing the promoter and the signal sequence of E. coli phoA gene, the structural gene for the periplasmic alkaline phosphatase. One of the most useful characteristics of this vector is the unique HindIII restriction site located just at the end of the phoA signal sequence. This restriction site was generated by oligonucleotide-directed site-specific mutagenesis without changing the amino acid sequence of the signal peptide. Any kind of foreign structural gene can be easily inserted into the HindIII site by using synthetic oligonucleotides to construct a hybrid gene which has neither an extra sequence nor a deletion between the phoA signal sequence and the foreign structural gene. Human alpha-interferon gene was inserted into this HindIII site. When this hybrid gene was expressed under the control of the phoA promoter region, a low but significant activity was recovered in the cold water wash of the cells after an osmotic shock procedure.

Alkaline Phosphatase↗
PubMed1985

Water and potassium ion absorption by deuterium oxide resistant winter rye seedlings.

The concentrations of deuterium oxide (D2O) in the root tissue water of winter rye seedlings equilibrated with the external D2O concentration within 30 min and in the shoot tissue water after 5-6 h. The equilibrated value in the root water was 87% and in the shoot water, 55% of the external concentration. The K+ absorption rate of the seedlings decreased from a value of 39 to 18 mumol g-1 h-1 when the D2O concentration was changed through a range from 0% to 97%. D2O suppressed the absorption of water and K+ by the seedlings. The higher the D2O concentration the greater the suppression, but it was less than with similarly treated rice plants. However, the process of D2O absorption by the seedlings was similar to that of rice seedlings (Envir. exp. Bot., 24, 369).

Absorption↗
PubMed1984

Length and structural effect of signal peptides derived from Bacillus subtilis alpha-amylase on secretion of Escherichia coli beta-lactamase in B. subtilis cells.

The precursor of Bacillus subtilis alpha-amylase contains an NH2-terminal extension of 41 amino acid residues as the signal sequence. The E. coli beta-lactamase structural gene was fused with the DNA for the promoter and signal sequence regions. Activity of beta-lactamase was expressed and more than 95% of the activity was secreted into the culture medium. DNA fragments coding for short signal sequences 28, 31, and 33 amino acids from the initiator Met were prepared and fused with the beta-lactamase structural gene. The sequences of 31 and 33 amino acid residues with Ala COOH-terminal amino acid were able to secrete active beta-lactamase from B. subtilis cells. However beta-lactamase was not secreted into the culture medium by the shorter signal sequence of 28 amino acid residues, which was not cleaved. Molecular weight analysis of the extracellular and cell-bound beta-lactamase suggested that the signal peptide of B. subtilis alpha-amylase was the first 31 amino acids from the initiator Met. The significance of these results was discussed in relation to the predicted secondary structure of the signal sequences.

Amino Acid Sequence↗
PubMed1984

A Bacillus subtilis secretion vector system derived from the B. subtilis alpha-amylase promoter and signal sequence region, and secretion of Escherichia coli beta-lactamase by the vector system.

A secretion vector system in Bacillus subtilis was constructed from the alpha-amylase promoter and signal sequence coding region of an alpha-amylase hyperproducing strain, B. subtilis NA64, and the major part of the plasmid pTUB4 which was derived from pUB110. When an Escherichia coli beta-lactamase gene, lacking its own promoter and signal sequence coding region, was introduced into the secretion vector system, beta-lactamase was expressed in B. subtilis. In addition, more than 95% of the enzyme synthesized was secreted into the culture medium via the secretion vector system. Secreted beta-lactamase crossreacted with rabbit antiserum raised against the E. coli enzyme.

Bacillus subtilis↗
PubMed1984

Discontinuous DNA replication of Drosophila melanogaster is primed by octaribonucleotide primer.

To investigate the precise structure of eucaryotic primer RNA made in vivo, short DNA chains isolated from nuclei of Drosophila melanogaster embryos were analyzed. Post-labeling of 5' ends of short DNA chains with polynucleotide kinase and [gamma-32P]ATP revealed that 7% of the DNA fragments were covalently linked with mono- to octaribonucleotide primers at their 5' ends. Octaribonucleotides, the major component (ca. 30%), formed the cap structure in the reaction with vaccinia guanylyltransferase and [alpha-32P]GTP, indicating that they were the intact primer RNA with tri- (or di-) phosphate termini, and the shorter ribooligomers were degradation intermediates. The intact primers started with purine (A/G ratio, 4:1), and the starting few ribonucleotide residues were rich in A.

Animals↗
PubMed1984

Hydrogen isotope effect on transport of potassium ion in rice seedlings equilibrated with deuterium oxide.

Potassium absorption by the root of rice seedlings, equilibrated with D2O before K+-absorption, decreased proportionally with D2O concentration and the translocation of the absorbed cations by roots to shoots decreased markedly above 40% D2O. An inhibition curve for the absorption process drawn as a relative rate vs. D2O concentration exhibits no threshold in concentration of D2O. While K+-absorption of the seedlings unequilibrated with D2O did not change up to 60% D2O and decreased above 80%. The concentration of D2O of root tissue water reached a balance with the external concentration within 30 min, but the concentration of shoot water required 5-6 h to balance with the external. The concentration of both root and shoot water did not coincide with the external. The suppressive effect of D2O on potassium absorption links not only to D/H ratio in root water but also in shoot water.

Adsorption↗