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B Tyler

Publications and source records attributed to B Tyler.

At least 37 records · Page 2Linked to original sources
PubMed1987

Mutations that create new promoters suppress the sigma 54 dependence of glnA transcription in Escherichia coli.

Escherichia coli rpoN mutants lack sigma 54 and are therefore unable to initiate the transcription of glnA at glnAp2, which is required for the production of a high intracellular concentration of glutamine synthetase. We have found that the dependence on sigma 54 can be overcome by mutations that have apparently created a new sigma 70-dependent promoter. The position -35 RNA polymerase contact site of this new promoter overlaps glnAp2. The initiation of transcription at the new promoter is inhibited by sigma 54-RNA polymerase even in the absence of nitrogen regulator I-phosphate, the activator required for the initiation of transcription at glnAp2. The results suggest that in cells growing with an excess of nitrogen and therefore lacking nitrogen regulator I-phosphate, sigma 54-RNA polymerase is bound at glnAp2.

Base Sequence↗
PubMed1986

Escherichia coli B/r leuK mutant lacking pseudouridine synthase I activity.

Escherichia coli B/r strain EB146 containing mutation leuK16 has elevated levels of enzymes involved in the synthesis of leucine, valine, isoleucine, histidine, and tryptophan (Brown et al., J. Bacteriol. 135:542-550, 1978). We show here that strain EB146 (leuK16) has properties that are similar to those of E. coli and Salmonella typhimurium hisT strains. In tRNA1Leu from both hisT and leuK strains, positions 39 and 41 are uridine residues rather than pseudouridine residues. Furthermore, in tRNA3Leu and tRNA4Leu from a leuK strain, uridine residues at positions 39 and 40, respectively, are unmodified. Pseudouridine synthase I activity is missing in extracts of strain EB146 (leuK16), and extracts of strain EB146 (leuK16) and of a hisT strain do not complement one another in vitro. Four phenotypes of strain EB146 (leuK16), leucine excretion, wrinkled colony morphology, and elevated levels of leu and his enzymes, are complemented by a plasmid having a 1.65-kilobase DNA fragment containing the E. coli K-12 hisT locus. These results indicate that either leuK codes for pseudouridine synthase I (and is thus a hisT locus in reality) or, less likely, it codes for a product that affects the synthesis or activity of pseudouridine synthase I.

Amino Acids↗
PubMed1983

Intracranial tuberculomas in a child: computed tomographic scan diagnosis and nonsurgical management.

Despite great strides in screening and detection of tuberculosis, urban areas still harbor many children with this disease. The case of a child with probable intracranial tuberculomas who was medically treated with antituberculous therapy and followed with frequent computed tomographic (CT) scans is reported. Despite older textbooks that espouse surgery as the treatment for intracranial tuberculomas, it is suggested that in the era of CT scanning, they should be treated the same as other focal CNS infections. Surgery should be reserved for medical failure or deteriorating conditions.

Antibiotics, Antitubercular↗
PubMed1982

Fine-structure deletion map and complementation analysis of the glnA-glnL-glnG region in Escherichia coli.

A total of 399 independent mutants of Escherichia coli were obtained which have point and insertion mutations in the glnA region. Mutants isolated included Gln- and Reg- strains (unable to utilize arginine as a nitrogen source). Mutations were mapped with 73 deletion-containing derivatives of a lambda gln phage. Complementation analysis was performed with lambda gln derivatives containing point mutations which conferred a Gln- or Reg- phenotype. Deletion mapping and complementation analysis assigned 104 mutations in 24 deletion intervals to glnA. Mutations in Reg- strains were assigned to two genes, glnL and glnG. glnL contained 131 mutations in 12 deletion intervals, and glnG contained 164 mutations in 10 deletion intervals. The gene order is glnA-glnL-glnG, transcribed from left to right. Polarity of insertion mutations indicates that glnL and glnG form from left to right. Polarity of insertion mutations indicates that glnL and glnG form an operon. Complementation analysis of glnA insertion mutations with glnL and glnG mutations showed polarity of glnA onto most glnL and glnG alleles, suggesting that transcription of glnA may proceed into the glnL-glnG operon. All mutations analyzed in glnA conferred a Gln- phenotype. However, we also found that over half of the Gln- strains isolated ater chemical mutagenesis contained point mutations in glnG. Mutants which synthesized a high level of glutamine synthetase in the presence of ammonia (GlnC phenotype) were selected as revertants of a strain with a Tn10 insertion in glnD and were mapped with chromosomal deletions. Results indicate that mutations in 12 and 15 examined strains clearly map outside of glnA, probably in glnL.

Chromosome Mapping↗
PubMed1982

Polarity in the glnA operon: suppression of the reg- phenotype by rho mutations.

To determine the ability of mutations in glnA, the gene for glutamine synthetase (GS), to regulate nitrogen assimilatory enzymes, we assayed histidase and GS in 34 glnA (Gln(-)) strains. Twenty-five glnA mutants were RegC, synthesizing high levels of histidase regardless of the availability of nitrogen, and nine were Reg(-), synthesizing low levels of histidase in medium containing either limiting or excess ammonia. rho mutations were introduced into strains containing glnA point mutations or insertions in glnA, glnL, glnG, or glnF. The Reg(-) phenotype of strains with glnA point mutations, but not those with glnA or glnF insertions, was altered by the presence of rho, suggesting that glnA (Reg(-)) mutations are polar and exert their phenotype by decreasing expression of glnL and glnG. Consistent with this view, no GS protein was detected by two-dimensional gel electrophoresis in glnA (Reg(-)) rho(+) or glnA (Reg(-)) rho double mutants, whereas GS protein was detected in cells of 10 of 11 glnA (RegC) strains. Since glnA (Reg(-)) rho double mutants synthesize constitutive levels of histidase, GS protein is not necessary for full expression of histidase. Mu d1 insertions in glnL, but not those in glnG, responded to the presence of a rho allele, presumably owing to elevated transcription into glnG from the Mu d1 prophage. Our results suggest that glnA (Reg(-)) alleles are polar mutations, and a rho-dependent termination site down-stream is postulated as the basis for the polar phenomenon. The data also indicate that, under some circumstances, a significant portion of glnL and glnG transcription is initiated at the glnA promoter.

DNA, Recombinant↗
PubMed1981

Regulation of glutamine synthetase activity by adenylylation in the Gram-positive bacterium Streptomyces cattleya.

The enzymatic activity of glutamine synthetase [GS; L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2] from the Gram-positive bacterium Streptomyces cattleya is regulated by covalent modification. In whole cells containing high levels of GS the addition of ammonium chloride leads to a rapid decline in GS activity. Crude extracts prepared from such ammonia-shocked cells had very low levels of GS activity as measured by biosynthetic and gamma-glutamyltransferase assays. Incubation of the crude extracts with snake venom phosphodiesterase restored GS activity. In cell extracts, GS was also inactivated by an ATP- and glutamine-dependent reaction. Radioactive labeling studies demonstrated the incorporation of an AmP moiety into GS protein upon modification. Our results suggest a covalent modification of GS in a Gram-positive bacterium. This modification appears to be adenylylation of the GS subunit similar to that found in the Gram-negative bacteria.

Adenine↗
PubMed1981

Organization of genes and spacers within the mouse immunoglobulin VH locus.

The germline organization of mouse immunoglobulin VH genes has been investigated using cloned VH sequences. Hybridization studies with VH probes from plasmacytomas HPC76 (H76), S107, HOPC1 (H1), and lymphoma ABLS-8 (A8) demonstrated that the VH locus contains at least three distinct VH gene families. Comparison with other results suggests a total of about 10 such families, and of the order of 160 germline VH genes. Nucleotide sequencing revealed that the H76 family includes anti-inulin VH sequences, and the S107 family is known to encode antiphosphorylcholine sequences. The three VH gene families studied were mapped in the order H76-S107-A8/H1-CH by determining which VH genes had been deleted from several plasmacytomas by VH rearrangement events. Nine genomic clones from athe H76 family, and one each from the S107 and A8/H1 families, were characterized; collectively they span 103 kilobases (kb). Two clones from the H76 family and one from the S107 family each bore a pair of VH genes separated by approximately 14 kb, suggesting that related VH genes in these families are clustered with a typical spacing of approximately 14 kb. No other VH genes were detected within the spacers, arguing against intermingling of different families. Within the VH76 family cluster, however, two closely homologous VH genes were shown not to be adjacent. While spacer sequences were strongly conserved in the A8/H1 family, the H76 family had minimal conservation and that was restricted to regions immediately surrounding the genes. Hence conservation of spacer sequences cannot be essential for VH gene function, nor for maintenance of a VN family. Spacers in both the H76 and S108 family contained small repeat elements, some of which behaved like mobile DNA sequences.

Animals↗
PubMed1980

Regulation of expression from the glnA promoter of Escherichia coli in the absence of glutamine synthetase.

One of the suspected regulators of glutamine synthetase [L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2] in enteric bacteria is glutamine synthetase itself. We isolated Escherichia coli strains carrying fusions of the beta-galactosidase structural gene to the promoter of the glutamine synthetase gene, with the aid of the Casadaban Mud1 (ApR, lac, cts62) phage. Some aspects of regulation were retained in haploid fusion strains despite the absence of glutamine synthetase, whereas other aspects required glutamine synthetase catalytic or regulatory activity or both. The direction of transcription of the glutamine synthetase gene was also determined.

Escherichia coli↗
PubMed1980

Purification of glutamine synthetase from a variety of bacteria.

We have developed two procedures which allow the very rapid purification of glutamine synthetase (GS) from a diverse variety of bacteria. The first procedure, based upon differential sedimentation, depends upon the association of GS with deoxyribonucleic acid in cell extracts. The second procedure, derived from the method of C. Gross et al (J. Bacteriol. 128:382-389, 1976) for purifying ribonucleic acid polymerase by polyethylene glycol (PEG) precipitation, enabled us to obtain high yields of GS from either small or large quantities of cells. We used the PEG procedure to purify GS from Klebsiella aerogenes, K. pneumoniae, Escherichia coli, Salmonella typhimurium, Rhizobium sp. strain 32H1, R. meliloti, Azotobacter vinelandii, Pseudomonas putida, Caulobacter crescentus, and Rhodopseudomonas capsulata. The purity of the GS obtained, judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was high, and in many instances only a single protein band was detected.

Azotobacter↗
PubMed1979

A new glnA-linked regulatory gene for glutamine synthetase in Escherichia coli.

Mutations in the glnA region of the Escherichia coli chromosome due to Mu prophage insertion result in two phenotypic classes. One class is Gln- and does not synthesize glutamine synthetase[L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2] under any growth condition. The other class produces a low level of glutamine synthetase under all growth conditions and is uncoupled from the regulatory effects of mutations in the glnF and glnD genes. Complementation analysis demonstrates that these two classes of insertions are in different cistrons. From these data we suggest that a regulatory gene, glnG, tightly linked to glnA, mediates both activation and repression of glutamine synthetase synthesis. An analysis of the evidence accumulated to date makes it unlikely that glnG is the only gene in the glnA region involved in the complex system of nitrogen regulation.

Bacteriophage mu↗
PubMed1979

Deletion mapping of the polA-metB region of the Escherichia coli chromosome.

A lambdacI857 prophage inserted into one of the genes of the rha locus was used to select deletions unambiguously ordering the markers polA-glnA-rha-pfkA-tpi-metBJF. Transduction with phage P1 indicates at least 70% linkage between glnA and polA. The order of the pfk and tpi markers is reversed from that previously published. Despite the relatively large distance separating the glnA and rha loci, deletions removing this entire region have no obvious phenotype. The isolation of Tn10 transposons integrated at different sites between rha and glnA greatly facilitated this work.

Chromosome Mapping↗
PubMed1979

Introduction of bacteriophage lambda into cells of Klebsiella aerogenes.

We have shown that a mutation in the cro gene of phage lambda greatly reduces zygotic induction. This observation has allowed us to move this phage on an episome into cells of Klebsiella aerogenes where it grows as well as in cells of Escherichia coli. This technique should allow the introduction of various derivatives of lambda into any organism which is able to receive deoxyribonucleic acid from E. coli.

Coliphages↗
PubMed1978

Regulation of glutamine synthetase formation in Escherichia coli: characterization of mutants lacking the uridylyltransferase.

A lambda phage (lambdaNK55) carrying the translocatable element Tn10, conferring tetracycline resistance (Tetr), has been utilized to isolate glutamine auxotrophs of Escherichia coli K-12. Such strains lack uridylyltransferase as a result of an insertion of the TN10 element in the glnD gene. The glnD::Tn10 insertion has been mapped at min 4 on the E. coli chromosome and 98% contransducible by phage P1 with dapD. A lambda transducing phage carrying the glnD gene has been identified. A glnD::Tn10 strain synthesizes highly adenylylated glutamine synthetase under all conditions of growth and fails to accumulate high levels of glutamine synthetase in response to nitrogen limitation. However, this strain, under nitrogen-limiting conditions, allows synthesis of 10 to 20 milliunits of biosynthetically active glutamine synthetase per mg of protein, which is sufficient to allow slow growth in the absence of glutamine. The GlnD phenotype in E. coli can be suppressed by the presence of mutations which increase the quantity of biosynthetically active glutamine synthetase.

Chromosome Mapping↗
PubMed1977

Regulation of glnA messinger ribonucleic acid synthesis in Klebsiella aerogenes.

We examined wild-type and mutant strains of Klebsiella aerogenes for the relative amounts of ribonucleic acid (RNA) hybridizing specifically to deoxyribonucleic acid from a transducing phage carrying glnAK, the structural gene for glutamine synthetase. Our data showed a positive correlation between the intracellular level of glutamine synthetase and the level of glnA messenger RNA; we were unable to detect glnA messinger RNA in strains devoid of glutamine synthetase protein. Therefore, it is possible that transcription of glnA is not regulated simply by repression mediated through the glutamine synthetase protein; rather, autogenous control in this system may involve activation of transcription. Our experiments also suggest that the promotor of the glnA gene is located at the rha proximal end of the gene.

Genes↗