scholarly journals Biosynthesis of soluble carnitine acetyltransferases from the yeast Candida tropicalis

1988 ◽  
Vol 253 (3) ◽  
pp. 845-849 ◽  
Author(s):  
B Kozulić ◽  
K Mosbach ◽  
F Meussdoerffer
Keyword(s):  
Continuous Culture ◽  
Candida Tropicalis ◽  
Growth Conditions ◽  
Proteolytic Processing ◽  
Carnitine Acetyltransferase ◽  
Acetyltransferase Activity ◽  
Tetrameric Form ◽  
In Cells

Soluble carnitine acetyltransferase from Candida tropicalis is synthesized as a 76 kDa precursor, which is monomeric and possesses no or very little carnitine acetyltransferase activity. Maturation of the enzyme begins with proteolytic processing of the 76 kDa precursor to 64 and 57 kDa subunits. The processed subunits subsequently associate into two kinds of active oligomers; the 57 kDa subunits are assembled into a tetramer and the 64 kDa subunits into an octamer. Formation of these oligomers depends apparently on growth conditions, since both oligomers were present in cells grown in continuous culture, but cells grown batchwise contained only the tetrameric form of carnitine acetyltransferase.

Efficient association of in vitro translation products with purified stable Candida tropicalis peroxisomes

1987 ◽  
Vol 7 (5) ◽  
pp. 1848-1855
Author(s):  
G M Small ◽  
T Imanaka ◽  
H Shio ◽  
P B Lazarow
Keyword(s):  
Candida Tropicalis ◽  
Proteolytic Processing ◽  
In Vitro Translation ◽  
Moderate Number ◽  
Brij 35 ◽  
Reticulocyte Lysate ◽  
Molecular Information ◽  
Acyl Coenzyme A

Newly synthesized peroxisomal proteins enter preexisting peroxisomes posttranslationally in vivo, generally without proteolytic processing. An efficient reconstitution of this process in vitro together with cloned DNAs for peroxisomal proteins would make possible investigation of the molecular information that targets proteins to peroxisomes. We have previously reported the isolation of clones for Candida tropicalis peroxisomal proteins; here we describe the association (and possible import) of peroxisomal proteins with peroxisomes in vitro. C. tropicalis was grown in a medium containing Brij 35, resulting in the induction of a moderate number of medium-sized peroxisomes. These peroxisomes, isolated in a sucrose gradient, had a catalase latency of 54% and were sufficiently stable to be concentrated and used in an import assay. The reticulocyte lysate translation products of total RNA from oleate-grown cells were incubated with the peroxisomes at 26 degrees C in the presence of 50 mM KCl, protease inhibitors, 0.5 M sucrose, 2.5 mM MOPS (morpholinepropanesulfonic acid) (pH 7.2), and 0.5 mM EDTA. Ten major translation products (which could be immunoprecipitated with antiserum against peroxisomal protein) became progressively associated with the peroxisomes during the first 30 min of incubation (some up to approximately 70%). These include acyl coenzyme A oxidase and the trifunctional protein hydratase-dehydrogenase-epimerase. This association did not occur at 4 degrees C nor did it occur if the peroxisomes were replaced with mitochondria.

scholarly journals Superoxide dismutase, catalase, and peroxidase in ammonium-grown and nitrogen-fixing Azospirillum brasilense

1984 ◽  
Vol 30 (10) ◽  
pp. 1222-1228 ◽  
Author(s):  
Richard W. Clara ◽  
Roger Knowles
Keyword(s):  
Superoxide Dismutase ◽  
Electron Acceptor ◽  
Azospirillum Brasilense ◽  
Polyacrylamide Gel Electrophoresis ◽  
Growth Conditions ◽  
Sod Activity ◽  
Batch Cultures ◽  
Physiological Conditions ◽  
Azospirillum Brasilense Sp7 ◽  
In Cells

Superoxide dismutase (SOD), catalase (CAT), and peroxidase (PER) activities were studied in ammonium-grown and N2-fixing batch cultures of Azospirillum brasilense Sp7. PER activity, as measured using o-dianisidine or 3,3′-diaminobenzidine as the H donor, was not significant in most growth conditions. SOD activity increased in response to higher O2 concentrations but was also present in cells grown anaerobically with nitrate [Formula: see text] or nitrous oxide (N2O) as electron acceptor. CAT activity increased at lower O2 concentrations and was highest in cells grown anaerobically with [Formula: see text] as electron acceptor. Polyacrylamide gel electrophoresis of cell-free extracts revealed only one band of SOD activity under each of the physiological conditions employed, compared with three for aerobically grown Escherichia coli K12. This band proved to be iron-containing SOD (FeSOD) on the basis of inhibitor sensitivity.

ABF1 is a phosphoprotein and plays a role in carbon source control of COX6 transcription in Saccharomyces cerevisiae

1992 ◽  
Vol 12 (9) ◽  
pp. 4197-4208
Author(s):  
S Silve ◽  
P R Rhode ◽  
B Coll ◽  
J Campbell ◽  
R O Poyton
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Carbon Source ◽  
Glucose Repression ◽  
Carbon Sources ◽  
Growth Conditions ◽  
Source Control ◽  
Specific Target Gene ◽  
Phosphorylation Pattern ◽  
Nonfermentable Carbon Source ◽  
In Cells

Previously, we have shown that the Saccharomyces cerevisiae DNA-binding protein ABF1 exists in at least two different electrophoretic forms (K. S. Sweder, P. R. Rhode, and J. L. Campbell, J. Biol. Chem. 263: 17270-17277, 1988). In this report, we show that these forms represent different states of phosphorylation of ABF1 and that at least four different phosphorylation states can be resolved electrophoretically. The ratios of these states to one another differ according to growth conditions and carbon source. Phosphorylation of ABF1 is therefore a regulated process. In nitrogen-starved cells or in cells grown on nonfermentable carbon sources (e.g., lactate), phosphorylated forms predominate, while in cells grown on fermentable carbon sources (e.g., glucose), dephosphorylated forms are enriched. The phosphorylation pattern is affected by mutations in the SNF1-SSN6 pathway, which is involved in glucose repression-depression. Whereas a functional SNF1 gene, which encodes a protein kinase, is not required for the phosphorylation of ABF1, a functional SSN6 gene is required for itsd ephosphorylation. The phosphorylation patterns that we have observed correlate with the regulation of a specific target gene, COX6, which encodes subunit VI of cytochrome c oxidase. Transcription of COX6 is repressed by growth in medium containing a fermentable carbon source and is derepressed by growth in medium containing a nonfermentable carbon source. COX6 repression-derepression is under the control of the SNF1-SSN6 pathway. This carbon source regulation is exerted through domain 1, a region of the upstream activation sequence UAS6 that binds ABF1 (J. D. Trawick, N. Kraut, F. Simon, and R. O. Poyton, Mol. Cell Biol. 12:2302-2314, 1992). We show that the greater the phosphorylation of ABF1, the greater the transcription of COX6. Furthermore, the ABF1-containing protein-DNA complexes formed at domain 1 differ according to the phosphorylation state of ABF1 and the carbon source on which the cells were grown. From these findings, we propose that the phosphorylation of ABF1 is involved in glucose repression-derepression of COX6 transcription.

scholarly journals Synthesis in vitro of precursor-type carnitine acetyltransferase with messenger RNA from Candida tropicalis

1984 ◽  
Vol 138 (3) ◽  
pp. 451-457 ◽  
Author(s):  
Mitsuyoshi UEDA ◽  
Atsuo TANAKA ◽  
Saburo HORIKAWA ◽  
Shosaku NUMA ◽  
Saburo FUKUI
Keyword(s):  
Candida Tropicalis ◽  
Messenger Rna ◽  
Carnitine Acetyltransferase

A mixed bacterial population in a continuous culture with and without kaolinite

1975 ◽  
Vol 21 (2) ◽  
pp. 173-180 ◽  
Author(s):  
R. Z. Maigetter ◽  
R. M. Pfister
Keyword(s):  
Continuous Culture ◽  
Bacterial Population ◽  
Fluorescent Antibody ◽  
Microscopic Analysis ◽  
Growth Conditions ◽  
Fluorescent Antibody Technique ◽  
Culture Vessel ◽  
Pseudomonas Sp ◽  
Culture Studies ◽  
Antibody Technique

Chromobacterium lividum and a Pseudomonas sp. were grown in pure and mixed continuous culture with and without the clay-mineral, kaolinite. Irrespective of the growth conditions, C. lividum adhered to the wall of the culture vessel whereas the Pseudomonas sp. showed no such tendency, at least visually. During mixed culture studies, the organism which was initially established in the culture dominated. The ratio between C. lividum and the Pseudomonas sp. was about 20:1 when C. lividum was first established and 1:2 when the Pseudomonas sp. was first grown. The indirect fluorescent antibody technique provided a rapid method for differentiating the mixed cultures when the bacterial concentration was sufficient for microscopic analysis. During both pure and mixed continuous culture studies, the addition of kaolinite reduced the C. lividum but not the Pseudomonas sp. population.

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