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S B Primrose

Publications and source records attributed to S B Primrose.

At least 19 recordsLinked to original sources
PubMed1986

The cloning and sequence analysis of the aspC and tyrB genes from Escherichia coli K12. Comparison of the primary structures of the aspartate aminotransferase and aromatic aminotransferase of E. coli with those of the pig aspartate aminotransferase isoenzymes.

In this paper we describe the cloning and sequence analysis of the tyrB and aspC genes from Escherichia coli K12, which encode the aromatic aminotransferase and aspartate aminotransferase respectively. The tyrB gene was isolated from a cosmid carrying the nearby dnaB gene, identified by its ability to complement a dnaB lesion. Deletion and linker insertion analysis located the tyrB gene to a 1.7-kilobase NruI-HindIII-digest fragment. Sequence analysis revealed a gene encoding a 43 000 Da polypeptide. The gene starts with a GTG codon and is closely followed by a structure resembling a rho independent terminator. The aspC gene was cloned by screening gene banks, prepared from a prototrophic E. coli K12 strain, for plasmids able to complement the aspC tyrB lesions in the aminotransferase-deficient strain HW225. Sub-cloning and deletion analysis located the aspC gene on a 1.8-kilobase HincII-StuI-digest fragment. Sequence analysis revealed the presence of a gene encoding a 43 000 Da protein, the sequence of which is identical with that previously obtained for the aspartate aminotransferase from E. coli B. Considerable overproduction of the two enzymes was demonstrated. We compared the deduced protein sequences with those of the pig mitochondrial and cytoplasmic aspartate aminotransferases. From the extensive homology observed we are able to propose that the two E. coli enzymes possess subunit structures, subunit interactions and coenzyme-binding and substrate-binding sites that are very similar both to each other and to those of the mammalian enzymes and therefore must also have very similar catalytic mechanisms. Comparison of the aspC and tyrB gene sequences reveals that they appear to have diverged as much as is possible within the constraints of functionality and codon usage.

Animals↗
PubMed1979

Biosynthesis of ethylene from methionine. Isolation of the putative intermediate 4-methylthio-2-oxobutanoate from culture fluids of bacteria and fungi.

Methods are described for identifying the 2,4-dinitrophenylhydrazones of 4-methylthio-2-oxobutanoate by means of t.l.c., n.m.r. and mass spectroscopy. By using these methods 4-methylthio-2-oxobutanoate, a putative intermediate in the biosynthesis of ethylene from methionine, has been identified in culture fluids of Aeromonas hydrophila B12E and a coryneform bacterium D7F grown in the presence of methionine. Relative to 4-methylthio-2-oxobutanoate, the yield of 3-(methylthio)propanal (methional) from the same cultures was less than 1%. Because 4-[2H]methylthio-2-oxobutanoate was obtained from cultures grown on [Me-2H]methionine, the 4-methylthio-2-oxobutanoate must be derived from methionine. By means of t.l.c. alone, 4-methylthio-2-oxobutanoate was identified in the culture fluids of a range of bacteria, the yeast Saccharomyces cerevisiae and the fungus Penicillium digitatum. A photochemical assay developed for 4-methylthio-2-oxobutanoate shows it to be a product of the metabolism of methionine by Escherichia, Pseudomonas, Bacillus, Acinetobacter, Aeromonas, Rhizobium and Corynebacterium species.

Bacteria↗
PubMed1979

Effect of glucose on polyol metabolism by Rhizobium trifolii.

The effect of glucose on polyol metabolism by Rhizobium trifolii was studied. Phenomena similar to catabolite repression and catabolite inhibition were observed. The catabolism of glucose to at least glucose 6-phosphate was required for the effects to be exerted.

Biological Transport↗
PubMed1978

Rapid purification of plasmid DNAs by hydroxyapatite chromatography.

A method is described for the rapid preparation of plasmid DNAs of molecular weight up to 14 X 10(6). This method involves the chromatography, at room temperature, of bacterial cleared lysates on hydroxyapatite in the presence of high concentrations of phosphate and urea. All detectable protein and RNA contamination of plasmid DNA is removed by this procedure and the conformation of the plasmid DNA is unaffected. Less than 0.5% chromosomal DNA is present in the purified preparation and even this can be removed if necessary by a simple extention of the procedure to include a heat-denaturation step. The method is extremely rapid and amenable to large-scale plasmid preparation; 5 mg ColE1 DNA have been purified within 40 min. The yield of plasmid DNA is similar to that obtained with the conventional dye-centrifugation technique, however the purity is greater.

Chromatography↗
PubMed1977

The effect of nalidixic acid on the cell cycle of synchronous Rhodopseudomonas palustris cultures.

The influence of the DNA synthesis inhibitor, nalidixic acid, on the properties of synchronous cultures of selected Rhodopseudomonas palustris swarmer cells was examined. There was little alteration in the changes in morphology, extinction, volume distribution and leucine incorporation up to bud development, and photosynthetic membrane lamellae were still synthesized de novo in the bud. However, there was no subsequent division, or flagellum or holdfast synthesis. Instead cells elongated by continued outgrowth of the abortive bud. Since DNA synthesis was also inhibited, this suggested a dependence of cell division, and flagellum and holdfast synthesis, on the completion of chromosome replication. By addition or removal of nalidixic acid at various times in the cell cycle, periods were demonstrated when the organism was insensitive to the antibiotic indicating that there was a pre-synthetic and post-synthetic gap in the pattern of DNA synthesis in R. palustris swarmers.

Cell Division↗
PubMed1977

Evaluation of the role of methional, 2-keto-4-methylthiobutyric acid and peroxidase in ethylene formation by Escherichia coli.

During growth of Escherichia coli strain SPA O in the presence of methionine, an intermediate accumulates in the medium. This intermediate reacts with 2,4-dinitrophenylhydrazine, and can be degraded to ethylene either enzymically or photochemically, the latter being stimulated by the addition of a flavin. The pH optimum for the photochemical degradation of this intermediate and 2-keto-4-methylthiobutyric acid (KMBA) is pH 3 whereas the optimum for methional is pH 6. The enzyme which converts the intermediate to ethylene also converts KMBA to ethylene and has many of the properties of a peroxidase including inhibition by catalase, cyanide, azide and anaerobiosis. The enzyme which synthesizes the intermediate is not known but requires oxygen and pyridoxal phosphate. A pathway for ethylene biosynthesis is proposed in which methionine is converted to KMBA which can be degraded either by peroxidase or in a flavin-mediated photochemical reaction. Its relevance to the properties of other ethylene-producing bacteria and to the proposed pathway of ethylene release by higher plants is discussed.

Aerobiosis↗
PubMed1976

Synchronous growth of Rhodopseudomonas palustris from the swarmer phase.

Rhodopseudomonas palustris was chosen as a model organism for studying bacterial differentiation. Synchronous populations selected by sucrose gradient centrifugation yielded more than 95% swarmer cells. The appearance and disappearance of cell morphological groupings and the doubling of cell numbers in cultures of such swarmer populations were very well defined. Cells were only motile for the first half of the division cycle, but motility was regained before division. Development gave rise to a distinct and characteristic pattern of extinction increase and particle volume distribution. The development of swarmers into mother cells and the dimorphic division of R. palustris are discussed as simple examples of differentiation.

Cell Count↗
PubMed1976

Formation of ethylene by Escherichia coli.

Escherichia coli strain SPA O converts methionine to ethylene by an inducible enzyme system. L-Cysteine, L-homocysteine, methionine derivatives and the sulphur-containing analogues of L-methionine also act as precursors of ethylene. Ethylene is produced by cell suspensions only in the presence of air; cell-free preparations can produce ethylene aerobically and anaerobically, but the extent to which they do so depends on the mode of culture growth. Light stimulates ethylene production by cell suspensions and its presence is essential for production by cell-free preparations. The kinetics of ethylene biogenesis and its pH and temperature optima suggest that ethylene is a secondary metabolite.

Aerobiosis↗