Continuous Culture Studies on the Biosynthesis of Alkaline Protease, Neutral Protease and  -Amylase by Bacillus subtilis NRRL-B3411

Heineken, F. G.; O’Connor, Richard J.

doi:10.1099/00221287-73-1-35

Cited by 76 publications

(27 citation statements)

References 14 publications

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“…This mechanism is not necessarily shared with any other genes, even including the gene for isocitrate dehydrogenase. For instance, amyE, like citB, is turned on at the end of the exponential phase in DSM medium (12), but the effects of addition of decoyinine on the transcription of these two genes are different. In the presence of glucose, the addition of decoyinine has no effect on expression of the amyE locus, unless the target of glucose repression has been inactivated by mutation (21)(22)(23).…”

Section: Discussionmentioning

confidence: 99%

A target for carbon source-dependent negative regulation of the citB promoter of Bacillus subtilis

1990

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Expression of the aconitase (citB) gene of Bacillus subtilis is subject to catabolite repression in cells grown in minimal media. In nutrient broth medium, citB expression is low in growing cells but is induced when cells enter sporulation. A 600-base-pair DNA fragment that extends from positions -400 through +200, relative to the transcription start site, was shown to include all of the cis-acting sequences necessary for catabolite repression and sporulation-associated regulation. This was demonstrated by fusing this DNA fragment to the Escherichia coli lacZ gene, integrating the fusion in the amyE locus of the B. subtilis chromosome, and measuring the regulation of expression of j8-galactosidase. By creating a series of deletions from either end of the 600-base-pair fragment, it was possible to define a target for catabolite repression; at least part of this target lies within the sequence between positions -84 and -68. DNA fragments that included positions -84 through +36, when carried on high-copy plasmids, caused derepression of aconitase synthesis, as if a negative regulator were being titrated. The same plasmids caused derepression of citrate synthase activity as well. Deletion of the sequence between positions -84 and -67 abolished this titration effect for both enzymes. Mutations that altered the target for catabolite repression also affected the inducibility of citB at the onset of sporulation, at least when sporulation was induced by the addition of decoyinine, an inhibitor of guanine nucleotide synthesis. When sporulation was induced by exhaustion of nutrient broth, there was no detectable difference in expression of citl-lacZ fusions whether or not they had the citB sequence from positions -84 to -67, suggesting that the mechanisms of regulation of citB in minimal medium and nutrient broth are different.Aconitase (EC 4.2.1.3; citrate [isocitrate] hydro-lyase), an enzyme of the Krebs cycle, is subject to several forms of regulation in Bacillus subtilis. In a minimal medium containing glucose and glutamine or glutamate, the citB gene, which encodes aconitase (5), is transcribed at a low level. Transcription is increased when glucose is replaced by a poorly metabolized carbon source such as citrate. It is also increased when the medium does not contain any compound rapidly convertible to 2-ketoglutarate (2-KG) (e.g., glutamine or glutamate) (26). A hint that this phenomenon might involve a negative regulator came from the observation that aconitase expression is partially derepressed in a cell that contains multiple copies of the citB promoter region (4).A second aspect of citB regulation is related to sporulation. Addition of decoyinine, a compound that induces sporulation in a defined medium by inhibiting the synthesis of guanine nucleotides (16), causes a rapid increase in citB transcription and aconitase activity (4,33). These two aspects of regulation may be linked in that induction of citB by decoyinine is associated with a decrease in 2-KG levels (4, 9, 32). It has been shown, but only at the le...

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Section: Discussionmentioning

confidence: 99%

A target for carbon source-dependent negative regulation of the citB promoter of Bacillus subtilis

1990

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“…The amyE gene, although not essential for sporulation, is also induced at the start of stationary phase and its expression is repressed by glucose (5,21,29). It is transcribed by the major vegetative form of RNA polymerase and its appearance is not affected by spoO mutations (W. Nicholson and G. Chambliss, personal communication).…”

Section: Discussionmentioning

confidence: 99%

Relationship between aconitase gene expression and sporulation in Bacillus subtilis

Dingman¹,

Rosenkrantz²,

Sonenshein³

1987

J Bacteriol

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The citB gene of Bacillus subtilis codes for aconitase (D. W. Dingman and A. L. Sonenshein,. By direct measurements of citB mRNA levels and by measurements of P-galatdase activity in a strain carrying a citE-lacZ fusion, we have examined the expression of citB during growth and sporulation. When cells were grown in nutrient broth sporulation medium, citB mRNA appeared in mid-to-lateexponential phase and disappeared by the second hour of sporulation. This timing corresponded closely to the kinetics of appearance of aconitase enzyme activity. Decoyinine, a compound that induces sporulation in a defined medium, caused a rapid simultaneous increase in aconitase activity and citB transcription. After decoyinine addition, the rate of increase in aconitase activity in a 2-ketoglutarate dehydrogenase (citk) mutant and in a citrate synthase (cit4) mutant was significantly less than in an isogenic wild-type strain. This is apparently due to a failure to deplete 2-ketoglutarate and accumulate citrate. These metabolites mit act as negative and positive effectors of citB expression, respectively. Mutations known to block sporulation at an early stage (spoOH and spoOB) had no appreciable effect on citB expression or aconitase activity. These results suggest that appearance of aconitase is stimulated by conditions that induce sporulation but is independent of certain gene products thought to act at an early stage of sporulation.Aconitase [EC 4.2.1.3; citrate (isocitrate) hydrolase], a tricarboxylic acid (TCA) cycle enzyme, appears to be subject to at least two forms of regulation in Bacillus subtilis. Catabolite repression of aconitase activity occurs whenever a rapidly metabolizable carbon source (e.g., glucose or mannose) is present in the medium (6). Catabolite repression is known to lower overall TCA cycle activity (6) and is thought to be a major regulating force for sporulation (20,33). For catabolite repression of aconitase to be complete, however, the medium must also contain a good source of 2-ketoglutarate (2-KG) (20,30). In the absence of a good carbon source, a high intracellular concentration of 2-KG has no detectable effect on aconitase activity (19 When cells of B. subtilis exhaust a complex medium and begin to sporulate, aconitase activity (which is at a very low level during growth) increases substantially 1 to 3 h after the onset of sporulation (14,16,36). This behavior might be taken to mean that aconitase is subject to sporulation control or that the signal that initiates sporulation also activates aconitase expression. Since aconitase activity in this case appears after the end of growth, it is possible that its regulation during sporulation is distinct from its regulation during growth in a defined medium. * Corresponding author. Mutants of B. subtilis lacking aconitase activity require glutamate (or glutamine) for growth and, after exhaustion of a complex medium, become blocked at stage 0 or I of sporulation (15,39). This again suggests a linkage between sporulation and aconitase expression. Three closel...

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“…A rapid loss of enzyme activity towards the end of the continuous culture experiments was observed; this might be due to an active degradation process. Recently, sharp declines in the enzyme activity of alkaline protease, neutral protease and s-amylase by Bacillus subtilis at later stages of continuous culturing have been reported due to spontaneous mutation to a less productive strain (Heineken and O'Connor, 1972).…”

Section: Discussionmentioning

confidence: 99%

Purification and properties of the alkaline phosphatase ofSerratia marcescens

Bhatti

1974

Arch. Microbiol.

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Abstract. A mutant strain of Serratia marcescens produces alkaline phosphatase in high yield. High concentrations of inorganic phosphate do not suppress formation of the enzyme and inhibition of its activity by inorganic phosphate is relatively low. The enzyme has been purified to homogeneity as determined by analytical ultracentrifugation and gel electrophoresis; it has a molecular weight of 100000. Guanidinium chloride reduces the apparent molecular weight about 50~ suggesting that the native enzyme is a dimer. Only one isoenzyme species is observed after electrophoresis. Specific activity is high (3553 units/mg protein); catalytic and spectroscopic properties are similar to those of alkaline phosphatase from other species. It is present to the extent of about 2 ~ of the cellular protein.

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Continuous Culture Studies on the Biosynthesis of Alkaline Protease, Neutral Protease and -Amylase by Bacillus subtilis NRRL-B3411

Cited by 76 publications

References 14 publications

A target for carbon source-dependent negative regulation of the citB promoter of Bacillus subtilis

A target for carbon source-dependent negative regulation of the citB promoter of Bacillus subtilis

Relationship between aconitase gene expression and sporulation in Bacillus subtilis

Purification and properties of the alkaline phosphatase ofSerratia marcescens

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