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Biomedical subjects

R B Helling

Publications and source records attributed to R B Helling.

At least 19 recordsLinked to original sources
PubMed1998

Pathway choice in glutamate synthesis in Escherichia coli.

Escherichia coli has two primary pathways for glutamate synthesis. The glutamine synthetase-glutamate synthase (GOGAT) pathway is essential for synthesis at low ammonium concentration and for regulation of the glutamine pool. The glutamate dehydrogenase (GDH) pathway is important during glucose-limited growth. It has been hypothesized that GDH is favored when the organism is stressed for energy, because the enzyme does not use ATP as does the GOGAT pathway. The results of competition experiments between the wild-type and a GDH-deficient mutant during glucose-limited growth in the presence of the nonmetabolizable glucose analog alpha-methylglucoside were consistent with the hypothesis. Enzyme measurements showed that levels of the enzymes of the glutamate pathways dropped as the organism passed from unrestricted to glucose-restricted growth. However, other conditions influencing pathway choice had no substantial effect on enzyme levels. Therefore, substrate availability and/or modulation of enzyme activity are likely to be major determinants of pathway choice in glutamate synthesis.

Ammonia↗
PubMed1995

icdB mutants of Escherichia coli.

icdB mutations map at 16 min, lead to the specific loss of citrate synthase, and are complemented by a prophage containing a gltA+ gene. Thus, they are allelic with gltA.

Chromosome Mapping↗
PubMed1994

Why does Escherichia coli have two primary pathways for synthesis of glutamate?

Escherichia coli has two primary pathways for glutamate synthetase-glutamate synthase pathway is known to be essential for synthesis at low ammonium concentrations and for regulation of the glutamine pool, but the necessity for glutamate dehydrogenase (GDH) has been uncertain. The results of competition experiments between the wild type and a GDH-deficient mutant during nutrient-limited growth and of direct enzyme measurements suggest that GDH is used in glutamate synthesis when the cell is limited for energy (and carbon) but ammonium and phosphate are present in excess, while the glutamine synthetase-glutamate synthase pathway is used when the cell is not under energy limitation. The use of alternative routes for glutamate synthesis implies that the energy cost of biosynthesis may be less when energy is limited than when energy is unlimited.

Escherichia coli↗
PubMed1992

Genetic changes accompanying increased fitness in evolving populations of Escherichia coli.

Two populations of Escherichia coli, each initiated with a single clone containing a derivative of the plasmid pBR322, were maintained for long periods in glucose-limited continuous culture. In both populations, after an extensive number of generations had elapsed, clones were isolated in which the transposon Tn3 from the plasmid had integrated into the bacterial chromosome. In both cases examined, the transpositions were shown to increase relative fitness approximately 6-7%, in the environment in which the populations were maintained. The loci of integration were mapped to approximately 13.2 min (population 1) and approximately 32.8 min (population 2).

Biological Evolution↗
PubMed1989

Processing of chloroplast ribosomal RNA transcripts in Euglena gracilis bacillaris.

The ribosomal RNA operons (rrn operons) of Euglena gracilis chloroplasts contain genes for (in order) 16S rRNA, tRNA(Ile), tRNA(Ala),23S rRNA and 5S rRNA. Major sites of cleavage of the primary rrn transcript were identified by Northern blot hybridization and S1-mapping. The presumptive termini of all of the mature products have now been identified. During initial processing in the chloroplast, the primary transcript is cleaved between the two tRNAs and between the 23S and 5S rRNAs so as to separate the sequences found in the different mature rRNAs. Subsequently the tRNAs are separated from the rRNAs, further trimming provides the remaining proper ends, and the 3'-ends of the tRNAs are added.

Animals↗
PubMed1988

Initiation of rrn transcription in chloroplasts of Euglena gracilis bacillaris.

The site of initiation of chloroplast rRNA synthesis was determined by S1-mapping and by sequencing primary rRNA transcripts specifically labeled at their 5'-end. Transcription initiates at a single site 53 nucleotides upstream of the 5'-end of the mature 16S rRNA under all growth conditions examined. The initiation site is within a DNA sequence that is highly homologous to and probably derived from a tRNA gene-region located elsewhere in the chloroplast genome. A nearly identical sequence (102 of 103 nucleotides) is present near the replication origin. The near identity of the two sequences suggests a common mode for control of transcription of the rRNA genes and initiation of chloroplast DNA replication. The related sequence in the tRNA gene-region does not appear to serve as a transcript initiation site.

Animals↗
PubMed1987

Evolution of Escherichia coli during growth in a constant environment.

Populations of Escherichia coli, initiated with a single clone and maintained for long periods in glucose-limited continuous culture, developed extensive polymorphisms. In one population, examined after 765 generations, two majority and two minority types were identified. Stable mixed populations were reestablished from the isolated strains. Factors involved in the development of this polymorphism included differences in the maximum specific growth rate and in the transport of glucose, and excretion of metabolites by some clones which were utilized by minority clones.

Adaptation, Physiological↗
PubMed1984

Sequence and evolution of the regions between thr rrn operons in the chloroplast genome of Euglena gracilis bacillaris.

The rRNA genes are arranged in three sequential operons preceded by a fourth partial operon. Part or all of a 1462 nucleotide sequence extending from within the 3'-end of the 23S rRNA gene, across the 5S rRNA gene and a presumptive transcription terminator, to within the first structural gene (for 16S rRNA) of the rrn operon was determined for each region between operons. Homologies of the 3'-end of the 23S rRNA gene with the 4.5S rRNA genes of higher plant chloroplasts, and of the 5S rRNA gene with other 5S rRNA genes were examined. The region preceding the 16S rRNA gene, which is expected to contain sites for initiation and regulation of rrn transcription, includes a 305 base-pair sequence with substantial homology with structural genes elsewhere in the chloroplast genome. The homologies suggest that this portion of the leader evolved from copies of parts of the structural genes which had been inserted before the 16S rRNA genes. Thus the chloroplast rrn leader may provide a unique opportunity to study how a regulatory sequence evolved from well-defined structural genes.

Base Sequence↗
PubMed1981

Cloning of mouse beta-casein gene sequences.

Casein messenger RNAs (mRNAcsn) were purified from lactating mammary glands of BALB/c mice and used as a starting material for cloning of casein gene sequences. Double-stranded casein cDNA (ds-cDNAcsn) was prepared and blunt-end ligated to HindIII-specific DNA linker molecules. After digestion with HindIII, the dsDNAcsn was inserted into the HindIII site of plasmid pBR322, using T4 DNA ligase. Escherichia coli strain RH202 was transformed with the hybrid plasmids, and transformants were selected for resistance to ampicillin. Electrophoresis of HindIII-digested hybrid plasmid DNAs, followed by Southern transfer and hybridization to [32P]cDNAcsn, revealed that one of the hybrid-plasmid-containing colonies, designated pCas51, contained a 400-bp insert which hybridized to the [32P]cDNAcsn. Purification of the individual casein mRNAs (mRNAcsn alpha, beta, and gamma) and solution hybridization of nick-translated insert DNA to each of these revealed that pCas51 contained sequences complementary primarily to mRNAcsn beta.

Animals↗
PubMed1981

The maintenance of Plasmid-containing organisms in populations of Escherichia coli.

Populations of Escherichia coli containing a small non-conjugative plasmid were grown in carbon-limited continuous culture. For all plasmids tested the presence of the plasmid lowered the growth rate of the host bacterium, and the proportion of plasmid-containing organisms in the total population declined initially. However, periodically, adaptive changes occurred in plasmid-containing organisms which increased their growth rate. This resulted in oscillations in the proportion of plasmid-containing organisms, and the delayed loss of the plasmid from the population.

Culture Media↗
PubMed1978

Acetate metabolism in Escherichia coli.

Levels of several intermediary metabolites were measured in cells grown in acetate medium in order to test the hypothesis that the glyoxylate cycle is repressed by phosphoenolpyruvate (PEP). Wild-type cells had less PEP than either isocitrate dehydrogenase - deficient cells (which had greater isocitrate lyase activity than the wild type) or isocitrate dehydrogenase - deficient, citrate synthase-deficient cells (which are poorly inducible). Thus induction of the glyoxylate cycle is more complicated than a simple function of PEP concentration. No correlation between enzyme activity and the level of oxaloacetate, pyruvate, or citrate was found either. Citrate was synthesized in citrate synthase-deficient mutants, possibly via citrate lyase.

Acetates↗
PubMed1977

Cloned ribosomal RNA genes from chloroplasts of Euglena gracilis.

Fragments of Euglena chloroplast DNA generated by endonuclease R-Eco RI were separated by agarose-gel electrophoresis into 24 distinct bands. At least five fragments contain sequences complementary to chloroplast ribosomal RNA, Most of the Eco RI fragments have been cloned in a plasmid of Escherichia coli. Three of the cloned fragments were shown to contain chloroplast ribosomal RNA sequences by DNA-RNA hybridization.

Chloroplasts↗
PubMed1976

Selection for citrate synthase deficiency in icd mutants of Escherichia coli.

Cultures of isocitrate dehydrogenase-deficient (icd) mutants were overgrown by double mutants (icd glt) lacking citrate synthase activity also. The icd mutants grew more slowly than wild-type cells or the double mutants because they accumulated an inhibitory metabolite (possibly citrate). Intracellular citrate levels were several hundred-fold higher in icd cells than in wild-type or icd glt cells. Final growth yields of the wild type and the icd mutant on limiting glucose were equivalent and greater than the growth yield of icd glt double mutants. The icd gene mapped between 60 and 74 min. icd mutants were resistant to nalidixic acid, but glt and icd glt mutants and wild-type cells were sensitive, indicating that resistance results from accumulation of isocitrate, citrate, or a derivative of these compounds.

Citrate (si)-Synthase↗