Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Mobilization of granulose in Clostridium pasteurianum. Purification and properties of granulose phosphorylase.

ABSTRACT: 1. The granulose of Clostridium pasteurianum ATCC 6013 is degraded when the organism is incubated in a medium containing no utilizable source of carbon and energy. 2. Mobilization of the polyglucan does not occur in the presence of exogenous glucose. 3. Breakdown of granulose is effected by a constitutively synthesized alpha-1,4-polyglucan phosphorylase. 4. Partial (530-fold) purification of this granulose phosphorylase was facilitated by its being loosely bound to the native granules of its substrate polyglucan. 5. The enzyme (pH optimum 6.4) was assayed both (a) in the degradative direction, K(m) for P(i)=2.2mm, and (b) in the synthetic direction, K(m) for glucose 1-phosphate=0.05mm. No requirement for bivalent cations was evidenced. 6. Granulose phosphorylase was inhibited by various nucleotide sugars; GDP-glucose, ADP-glucose (K(i)=20mum) and UDP-glucose (K(i)=60mum) were particularly potent competitive inhibitors. ATP, NADP(+) and NADPH (at 1mm) were less effective inhibitors, whereas AMP was slightly stimulatory. 7. It would appear that granulose mobilization is favoured under conditions of low adenylate energy charge, but is prevented under conditions of ;glucose excess' chiefly by ADP-glucose-mediated inhibition of granulose phosphorylase.

SUBMITTER: Robson RL

PROVIDER: S-EPMC1168529 | biostudies-other | 1974 Dec

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Purification and properties of threonine aldolase from Clostridium pasteurianum.

Project description:1. Threonine aldolase was purified about 200-fold in 10% yield from Clostridium pasteurianum and its properties were examined. The final preparation gave three bands after ionophoresis on polyacrylamide gel. 2. The purified enzyme was shown to produce glycine and acetaldehyde in stoicheiometric amounts from threonine. The reverse reaction was demonstrated qualitatively. 3. The enzyme has a broad pH optimum at 6.5-7.0. 4. The enzyme is highly specific for l-threonine. 5. The enzyme is completely inhibited by 1mm concentrations of hydroxylamine and semicarbazide. Activity is decreased to 20% of the original by treatment with cysteine plus mercaptoethanol; most of the loss is regained on incubation with pyridoxal phosphate. It is concluded that pyridoxal phosphate is a prosthetic group. 6. The relationship between velocity and substrate concentration is atypical but indicates a K(m) value of 0.42mm. 7. The enzyme was demonstrated in several other strictly anaerobic bacteria.

| S-EPMC1178963 | biostudies-other

Clostridium pasteurianum F1Fo ATP synthase: operon, composition, and some properties.

Project description:The atp operon encoding F1Fo ATP synthase in the fermentative obligate anaerobic bacterium Clostridium pasteurianum was sequenced. It consisted of nine genes arranged in the order atpI(i), atpB(a), atpE(c), atpF(b), atpH(delta), atpA(alpha), atpG(gamma), atpD(beta), and atpC(epsilon), which was identical to that found in many bacteria. Reverse transcription-PCR confirmed the presence of the transcripts of all nine genes. The amount of ATPase activity in the membranes of C. pasteurianum was low compared to what has been found in many other bacteria. The F1Fo complexes solubilized from membranes of C. pasteurianum and Escherichia coli had similar masses, suggesting similar compositions for the F1Fo complexes from the two bacteria. Western blotting experiments with antibodies raised against the purified subunits of F1Fo detected the presence of eight subunits, alpha, beta, gamma, delta, epsilon, a, b, and c, in the F1Fo complex from C. pasteurianum. The F1Fo complex from C. pasteurianum was activated by thiocyanate, cyanate, or sulfhydryl compounds; inhibited by sulfite, bisulfite, or bicarbonate; and had tolerance to inhibition by dicyclohexylcarbodiimide. The target of thiol activation of the F1Fo complex from C. pasteurianum was F1. Thiocyanate and sulfite were noncompetitive with respect to substrate Mg ATP but competitive with respect to each other. The F1 and Fo parts of the F1Fo complexes from C. pasteurianum and E. coli bound to each other, but the hybrid F1Fo complexes were not functionally active.

| S-EPMC193768 | biostudies-literature

Clostridium pasteurianum

Project description:Nitrogen-fixing soil bacterium

| PRJNA35233 | ENA

Clostridium pasteurianum

Project description:Clostridium pasteurianum Genome sequencing

| PRJNA299496 | ENA

Closed Genome Sequence of Clostridium pasteurianum ATCC 6013.

Project description:We report here the closed genome of Clostridium pasteurianum ATCC 6013, a saccharolytic, nitrogen-fixing, and spore-forming Gram-positive obligate anaerobe. The organism is of biotechnological interest due to the production of solvents (butanol and 1,3-propanediol) but can be associated with food spoilage. The genome comprises a total of 4,351,223 bp.

| S-EPMC4335343 | biostudies-literature

Mechanism of proton transfer in [FeFe]-hydrogenase from Clostridium pasteurianum.

Project description:[FeFe]-Hydrogenases are complex metalloproteins that catalyze the reversible reduction of protons to molecular hydrogen utilizing a unique diiron subcluster bridged to a [4Fe4S] subcluster. Extensive studies have concentrated on the nature and catalytic activity of the active site, yet relatively little information is available concerning the mechanism of proton transport that is required for this activity. Previously, structural characterization of [FeFe]-hydrogenase from Clostridium pasteurianum indicated a potential proton transport pathway involving four residues (Cys-299, Glu-279, Ser-319, and Glu-282) that connect the active site to the enzyme surface. Here, we demonstrate that substitution of any of these residues resulted in a drastic reduction in hydrogenase activity relative to the native enzyme, supporting the importance of these residues in catalysis. Inhibition studies of native and amino acid-substituted enzymes revealed that Zn(2+) specifically blocked proton transfer by binding to Glu-282, confirming the role of this residue in the identified pathway. In addition, all four of these residues are strictly conserved, suggesting that they may form a proton transport pathway that is common to all [FeFe]-hydrogenases.

| S-EPMC3207428 | biostudies-literature

Clostridium pasteurianum BC1

Project description:Nitrogen-fixing bacterium and potential bioremediator.

| PRJNA80 | ENA

Towards improved butanol production through targeted genetic modification of Clostridium pasteurianum.

Project description:Declining fossil fuel reserves, coupled with environmental concerns over their continued extraction and exploitation have led to strenuous efforts to identify renewable routes to energy and fuels. One attractive option is to convert glycerol, a by-product of the biodiesel industry, into n-butanol, an industrially important chemical and potential liquid transportation fuel, using Clostridium pasteurianum. Under certain growth conditions this Clostridium species has been shown to predominantly produce n-butanol, together with ethanol and 1,3-propanediol, when grown on glycerol. Further increases in the yields of n-butanol produced by C. pasteurianum could be accomplished through rational metabolic engineering of the strain. Accordingly, in the current report we have developed and exemplified a robust tool kit for the metabolic engineering of C. pasteurianum and used the system to make the first reported in-frame deletion mutants of pivotal genes involved in solvent production, namely hydA (hydrogenase), rex (Redox response regulator) and dhaBCE (glycerol dehydratase). We were, for the first time in C. pasteurianum, able to eliminate 1,3-propanediol synthesis and demonstrate its production was essential for growth on glycerol as a carbon source. Inactivation of both rex and hydA resulted in increased n-butanol titres, representing the first steps towards improving the utilisation of C. pasteurianum as a chassis for the industrial production of this important chemical.

| S-EPMC5367854 | biostudies-literature

The biosynthesis of granulose by Clostridium pasteurianum.

Project description:1. Mutant strains of Clostridium pasteurianum were obtained, which are unable to synthesize granulose (an intracellularly accumulated amylopectin-like alpha-polyglucan). 2. These mutants lacked either (a) ADP-glucose pyrophosphorylase (EC 2.7.7.27), or (b) granulose synthase (i.e. ADP-glucose-alpha-1,4-glucan glucosyltransferase, EC 2.4.1.21). 3. Although both of these enzymes were constitutively synthesized by the wild-type organism, massive deposition of granulose in a sporulating culture coincided with a threefold increase in the specific activity of ADP-glucose pyrophosphorylase. 4. The soluble ADP-glucose pyrophosphorylase was partially purified (33-fold). Its ATP-saturation curve was not sigmoidal and its activity was not enhanced by phosphorylated intermediates of glycolysis, pyruvate, NAD(P)H or pyridoxal 5'-phosphate. ADP at relatively high concentrations acted as a competitive inhibitor (K(i)=19mm). 5. The dependence of granulose synthase on a suitable polyglucan primer was demonstrated by using enzyme obtained from a granulose-free mutant strain (lacking ADP-glucose pyrophosphorylase). 6. Partial purification of granulose synthase from wild-type strains was facilitated by its being bound to the native particles of granulose. No activator was discovered, but ADP, AMP and pyridoxal 5'-phosphate were competitive inhibitors, ADP being most effective (K(i) about 0.2mm). 7. It would appear that the synthesis of granulose in Cl. pasteurianum is not subject to the positive, fine control that is a feature of glycogen biosynthesis in most bacteria.

| S-EPMC1168528 | biostudies-other

Biosynthesis of amino acids in Clostridium pasteurianum.

Project description:1. Clostridium pasteurianum was grown on a synthetic medium with the following carbon sources: (a) (14)C-labelled glucose, alone or with unlabelled aspartate or glutamate, or (b) unlabelled glucose plus (14)C-labelled aspartate, glutamate, threonine, serine or glycine. The incorporation of (14)C into the amino acids of the cell protein was examined. 2. In both series of experiments carbon from exogenous glutamate was incorporated into proline and arginine; carbon from aspartate was incorporated into glutamate, proline, arginine, lysine, methionine, threonine, isoleucine, glycine and serine. Incorporations from the other exogenous amino acids indicated the metabolic sequence: aspartate --> threonine --> glycine right harpoon over left harpoon serine. 3. The following activities were demonstrated in cell-free extracts of the organism: (a) the formation of aspartate by carboxylation of phosphoenolpyruvate or pyruvate, followed by transamination; (b) the individual reactions of the tricarboxylic acid route to 2-oxoglutarate from oxaloacetate; glutamate dehydrogenase was not detected; (c) the conversion of aspartate into threonine via homoserine; (d) the conversion of threonine into glycine by a constitutive threonine aldolase; (e) serine transaminase, phosphoserine transaminase, glycerate dehydrogenase and phosphoglycerate dehydrogenase. This last activity was abnormally high. 4. The combined evidence indicates that in C. pasteurianum the biosynthetic role of aspartate and glutamate is generally similar to that in aerobic and facultatively aerobic organisms, but that glycine is synthesized from glucose via aspartate and threonine.

| S-EPMC1178962 | biostudies-other

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data