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Biomedical subjects

H Paulus

Publications and source records attributed to H Paulus.

At least 91 records · Page 5Linked to original sources
PubMed1978

Isolation and characterization of polyadenylate-containing RNA from Bacillus brevis.

A substantial fraction (30--40%) of pulse-labeled RNA from exponentially growing cells of Bacillus brevis contains polyadenylate sequences, as measured by adsorption to oligo(dT)-cellulose. The weight-average length of poly(A) tracts obtained after digestion with pancreatic and T1 ribonucleases is 60 nucleotide residues. Susceptibility to degradation by snake venom phosphodiesterase after ribonuclease degradation indicates that the poly(A) sequences are located near the 3' ends of the RNA chains, but that in 40% of the material at least one internal pyrimidine nucleotide residue intervenes between the poly(A) tract and the 3'-hydroxyl terminus. These pyrimidine nucleotides consist of 65% cytidylate and 35% uridylate residues. In the remaining RNA chains, the poly(A) sequence is directly at the 3'-terminus, but the possibility cannot be excluded that a small fraction of this material may contain a 3'-hydroxyl terminal guanylate residue. The weight-average sedimentation coefficient of poly(A)-containing RNA is 12.5 S, corresponding to a polynucleotide chain length of 800--900 residues. This is in a size range expected for messenger RNA, a possibility which is also supported by the observation that pulse-labeled RNA has a considerably higher poly(A) content than long-term labeled RNA.

Bacillus↗
PubMed1977

Biological function of gramicidin: studies on gramicidin-negative mutants.

By the use of a rapid radioautographic screening procedure, two mutants of Bacillus brevis ATCC 8185 that have lost the ability to produce gramicidin have been isolated. These mutants produced normal levels of tyrocidine and sporulated at the same frequency as the parent strain. Their spores, however, were more heat-sensitive and had a reduced dipicolinic acid content. Gramicidin-producing revertants occurred at a relatively high frequency among the survivors of prolonged heat treatment and had also regained the ability to produce heat-resistant spores. A normal sport phenotype could also be restored by the addition of gramicidin to cultures of the mutant strain at the end of the exponential growth. On the other hand, the addition of dipicolinic acid could not cure the spore defect. These results provide strong evidence that the inability to produce gramicidin is directly responsible for the observed spore defects. Indeed, they unambiguously demonstrate a function of a peptide antibiotic in bacterial sporulation. The possibility that this function consists of the regulation of transcription during the transition from growth to sporulation is discussed.

Bacillus↗
PubMed1977

Biological function of gramicidin: selective inhibition of RNA polymerase.

This paper describes a novel biochemical effect of gramicidin, a class of peptide antibiotics produced by Bacillus brevis during the transition from vegetative growth to sporulation. Gramicidin inhibits RNA synthesis by purified RNA polymerase (nucleosidetriphosphate:RNA nucleotidyl-transferase, EC 2.7.7.6) by interfering with the binding of RNA polymerase to DNA. This effect seems to involve the destabilization of the "open" RNA polymerase-DNA complex, a mode of action consistent with the control of promoter selection. Selectivity in the inhibition of RNA synthesis by gramicidin is observed when transcription is partially blocked by low levels of actinomycin D. Since the inhibition of RNA synthesis by gramicidin is obtained in a highly purified system devoid of membranes, it must be distinct from the ionophoretic activity of the antibiotic. It is possible that this new mode of action reflects the function of gramicidin during bacterial sporulation.

Bacillus↗
PubMed1976

Indomethacin serum concentrations in man. Effects of dosage, food, and antacid.

In the use of anti-inflammatory compounds, sustained serum levels are thought to be related to drug efficacy. This study shows that frequent clinical administration of indomethacin can result in sustained serum levels of the drug and that food and antacid may have important modifying effects on serum indomethacin concentrations. After oral ingestion by fasting subjects, indomethacin rapidly appeared in the serum, usually reaching peak concentrations in 30 to 90 minutes. Food delayed and decreased the mean peak level; antacid delayed the peak and slightly enhanced subsequent concentrations. With multiple dose schedules plateau levels were reached after 24 hours. When a total daily dose of 150 mg was given as 25 mg every 4 hours peak concentrations were the same but fluctuations were smaller and average concentrations were higher than with a dosage of 50 mg every 8 hours.

Administration, Oral↗
PubMed1975

Purification and properties of homoserine transacetylase from Bacillus polymyxa.

Homoserine transacetylase (EC 2.3.1.31), the first enzyme of methionine biosynthesis, has been purified to near homogeneity from extracts of a methionine auxotroph of Bacillus polymyxa. The enzyme is subject to rapid irreversible inactivation. Its half-life at 0 degrees is 15 min and much less at higher temperatures, but ethylene glycol affords some protection. In addition, Zn2+ reversibly inhibits the enzyme with a K-I of 3 muM. The enzyme has a molecular weight of about 40,000 and consists of a single polypeptide chain. Besides catalyzing the acetyl transfer from acetyl-CoA to L-homoserine, homoserine transacetylase promotes a homoserine-O-acetylhomoserine exchange reaction in the absence of CoA, suggesting the formation of an acetyl-enzyme intermediate. The results of kinetic studies are consistent with a ping-pong mechanism. Homoserine transacetylase is subject to multivalent feedback inhibition by L-methionine and S-adenosylmethionine. Analysis of the inhibition data and specificity studies suggest that the inhibitors bind to separate sites on the enzyme which are distinct from the active site. Inhibition is competitive with respect to both substrates, and the saturation curves for the inhibitors, as well as substrate saturation curves in the absence or presence of the inhibitors, are hyperbolic. The absence of cooperativity is, in fact, a property which would be expected in a monomeric allosteric enzyme such as homoserine transacetylase.

Acetyl Coenzyme A↗
PubMed1975

Regulation of homoserine transacetylase in whole cells of Bacillus polymyxa.

The levels of homoserine transacetylase (EC 2.3.1.31) in Bacillus polymyxa grown in minimal medium can vary over a 40-fold range, depending on whether methionine limits growth or is present in excess. This suggests that the synthesis of the enzyme is under control by methionine or one of its metabolites. The stability of homoserine transacetylase in growing cells was measured after repression of further synthesis by the addition of methionine. At 30 degrees, the enzyme was stable for 2 hours, whereas at 37 degrees it decayed with a half-life of 40 min. This contrasts with the striking instability in cell-free extracts described in the preceding paper (Wyman, A., and Paulus, H. (1975) J. Biol. Chem. 250, 3897-3903). The properties of homoserine transacetylase were also studied in cells of B. polymyxa that had been made permeable to small molecules by treatment with toluene. They differed in two important respects from those of the enzyme in cell-free extracts described in the preceding paper: the enzyme was relatively stable, with a half-life of 15 min at 37 degrees, and responded in a sigmoid manner to increasing concentrations of the inhibitors L-methionine and S-adenosylmethionine. These observations suggest that homoserine transacetylase is an oligomeric protein within the bacterial cell but dissociates into monomers in cell-free extracts. When B. polymyxa was transferred at 39 degrees from a rich medium to one without amino acids, growth resumed only very slowly. The growth lag after shift-down was not observed at 37 degrees or in the presence of methionine or cystathionine. This phenomenon appears to be due to a need for derepression of homoserine transacetylase upon shift-down which is thwarted at 39 degrees by the rapid thermal inactivation of the enzyme. A possible physiological function of the striking thermolability of the first enzyme in methionine biosynthesis is discussed.

Acetyltransferases↗
PubMed1975

A guanosine 3':5'-monophosphate-sensitive nuclease from Bacillus brevis.

In toluene-treated cells of Bacillus brevis, newly synthesized RNA is rapidly degraded in a reaction that is inhibited by cyclic guanosine 3':5'-monophosphate (cGMP) and by 1,10-phenanthroline. This appears to be due to a ribonuclease found in cell-free extracts of B. brevis which is inhibited by cGMP and related compounds as well as by 1,10-phenanthroline. The cGMP-sensitive nuclease hydrolyzes synthetic polynucleotides, yielding nucleoside 5'-monophosphates as the sole products, even during the early stages of hydrolysis. Synthetic polynucleotides terminated by a 3'-phosphate are resistant to hydrolysis. While with 3'-hydrolysis of the polymer. The enzyme is therefore an exonuclease that degrades polynucleotides from the 3' end to product 5'-mononucleotides. It also acts on denatured but not on native DNA. Activity is greatest in the presence of Mn2+ and is not affected by the presence of monovalent cations. 1,10-Phenanthroline, but not 1,7-phenanthroline, inhibits the nuclease even when Mn2+ is present in excess. The inhibition of the enzyme by cGMP is noncompetitive, and cGMP itself is not hydrolyzed. The sensitivity of the nuclease to inhibition depends strikingly on the nature of the substrate and is lost when the enzyme is assayed at high pH. These observations suggest that cGMP inhibits the nuclease by combining with an allosteric site on the enzyme. Although cGMP was found to be the most effective inhibitor, other nucleoside 3':5'-monophosphates and derivatives of 5'-GMP can also inhibit the nuclease. Since measurements of cGMP in B. brevis have not revealed detectable amounts (less than 5 times 10-8 M), the substance that modulates the activity of the nuclease under physiological conditions remains to be identified.

Bacillus↗
PubMed1972

Nucleotide-dependent inactivation of RNA polymerase from Bacillus brevis.

RNA polymerase has been purified from vegetative cells of Bacillus brevis and resolved into "core" enzyme and sigma factor. The purified enzyme is rapidly inactivated by incubation at low temperatures in the presence of 1-2 mM ATP, dATP, or NAD(+), while other nucleotides at this concentration have little or no effect. Inactivation is not accompanied by the incorporation of an adenylyl or phosphoryl moiety into RNA polymerase; nevertheless, it is essentially irreversible. DNA, high concentrations of glycerol, as well as low concentrations (1 mM) of orthophosphate protect RNA polymerase from the nucleotide-dependent inactivation.A similar inactivation of RNA polymerase in the presence of ATP is observed with crude preparations from Bacillus subtilis and Bacillus polymyxa. This phenomenon may represent a novel mode of regulation of transcription that does not involve a covalent modification of RNA polymerase or its interaction with other protein factors, but rather is due to a structural transition to an inactive form induced by small molecules.

Adenosine Diphosphate↗