scholarly journals The pathway of glycollate utilization in Chlorella pyrenoidosa

1970 ◽  
Vol 117 (5) ◽  
pp. 929-937 ◽  
Author(s):  
J. M. Lord ◽  
M. J. Merrett
Keyword(s):  
Soluble Fraction ◽  
Tricarboxylic Acid Cycle ◽  
Experimental Period ◽  
Chlorella Pyrenoidosa ◽  
Total Radioactivity ◽  
Tricarboxylic Acid ◽  
Short Term ◽  
Acid Cycle ◽  
Metabolic Sequence ◽  
Specific Activities

1. Exogenous glycollate was rapidly metabolized in both the light and the dark by photoautotrophically grown Chlorella pyrenoidosa. 2. The incorporation of 14C from [1-14C]glycollate by these cells was inhibited by the tricarboxylic acid-cycle inhibitors monofluoroacetate, diethylmalonate and arsenite, and also by α-hydroxypyrid-2-ylmethanesulphonate and isonicotinylhydrazine. 3. Short-term kinetic experiments showed over 80% of the total 14C present in the soluble fraction from the cells to be in glycine and serine after 10s. This percentage decreased with time whereas the percentage radioactivity in glycerate increased for up to 30s then remained steady. The percentage of the total radioactivity present in citrate increased over the experimental period. Malate was the only other tricarboxylic acid-cycle intermediate to become labelled. 4. The kinetic and inhibitor experiments supported the following pathway of glycollate incorporation: glycollate → glyoxylate → glycine → serine → hydroxypyruvate → glycerate → 3-phosphoglycerate → 2-phosphoglycerate → phosphoenolpyruvate → pyruvate → acetyl-CoA. 5. The specific activities of the enzymes catalysing this metabolic sequence in cell-free extracts were great enough to account for the observed rate of glycollate metabolism of 0.25μmol/h per mg dry wt. of cells in the light.

scholarly journals A Soluble NADH-Dependent Fumarate Reductase in the Reductive Tricarboxylic Acid Cycle of Hydrogenobacter thermophilus TK-6

2008 ◽  
Vol 190 (21) ◽  
pp. 7170-7177 ◽  
Author(s):  
Akane Miura ◽  
Masafumi Kameya ◽  
Hiroyuki Arai ◽  
Masaharu Ishii ◽  
Yasuo Igarashi
Keyword(s):  
Soluble Fraction ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Fumarate Reductase ◽  
High Sequence Identity ◽  
Catalytic Subunits ◽  
Acid Cycle ◽  
Genes Encoding ◽  
Hydrogenobacter Thermophilus ◽  
Reductive Tricarboxylic Acid Cycle

ABSTRACT Fumarate reductase (FRD) is an enzyme that reduces fumarate to succinate. In many organisms, it is bound to the membrane and uses electron donors such as quinol. In this study, an FRD from a thermophilic chemolithoautotrophic bacterium, Hydrogenobacter thermophilus TK-6, was purified and characterized. FRD activity using NADH as an electron donor was not detected in the membrane fraction but was found in the soluble fraction. The purified enzyme was demonstrated to be a novel type of FRD, consisting of five subunits. One subunit showed high sequence identity to the catalytic subunits of known FRDs. Although the genes of typical FRDs are assembled in a cluster, the five genes encoding the H. thermophilus FRD were distant from each other in the genome. Furthermore, phylogenetic analysis showed that the H. thermophilus FRD was located in a distinct position from those of known soluble FRDs. This is the first report of a soluble NADH-dependent FRD in Bacteria and of the purification of a FRD that operates in the reductive tricarboxylic acid cycle.

scholarly journals Effect of ethanol on lipid metabolism in cultured hepatocytes

1985 ◽  
Vol 228 (3) ◽  
pp. 673-681 ◽  
Author(s):  
N Grunnet ◽  
J Kondrup ◽  
J Dich
Keyword(s):  
Fatty Acid ◽  
Tricarboxylic Acid Cycle ◽  
Fatty Acid Uptake ◽  
Tricarboxylic Acid ◽  
Acid Uptake ◽  
Short Term ◽  
Acid Cycle ◽  
Body Production ◽  
Rate Of Accumulation

Isolated rat hepatocytes were cultured in a modified HI-WO/BA medium for 16 h. In the following 24 h oleate or oleate plus ethanol was added to the medium. After this period the medium was changed again and the cultures were further incubated with [1-14C]oleate alone or with [1-14C]oleate plus ethanol for 6 h. This allowed a comparison of effects of short-term (6 h) and long-term (24 + 6 h) exposure to ethanol on fatty acid metabolism. The increased intracellular accumulation of triacylglycerol in the presence of ethanol was quantitatively accounted for by increased fatty acid uptake, by decreased fatty acid oxidation in the tricarboxylic acid cycle and by decreased VLDL (very-low-density lipoprotein)-triacylglycerol secretion. Ketone-body production was not affected. After short-term exposure the rate of accumulation of triacylglycerol was increased by 50%. This increase was accounted for by increased fatty acid uptake (44%), decreased tricarboxylic acid-cycle activity (49%) and decreased VLDL-triacylglycerol secretion (7%). After long-term exposure, the rate of accumulation of triacylglycerol was increased by 74%. This increase was accounted for by increased fatty acid uptake (34%), decreased tricarboxylic acid-cycle activity (34%) and decreased VLDL-triacylglycerol secretion (32%). The larger increase in accumulation of triacylglycerol after long-term exposure to ethanol was entirely accounted for by increased inhibition of secretion of VLDL-triacylglycerol. The biochemical mechanisms underlying the observations are discussed.

Effect of oxygen supply during growth on the production of cytochromes, enzymes, and acid end products by Haemophilus parasuis

1992 ◽  
Vol 38 (9) ◽  
pp. 958-968
Author(s):  
P. G. Martin ◽  
D. F. Niven
Keyword(s):  
Phosphoenolpyruvate Carboxylase ◽  
Oxygen Supply ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Haemophilus Parasuis ◽  
Acid Cycle ◽  
End Products ◽  
Specific Activities ◽  
Effect Of Oxygen ◽  
Reductive Pathway

Haemophilus parasuis, grown under conditions of high aeration, was found to lack a tricarboxylic acid cycle but to possess phosphoenolpyruvate carboxylase and a reductive pathway leading to the production of succinate. Such organisms contained approximately equal quantities of b-, c-, and d-type cytochromes and excreted acetate. When the oxygen supply for growth was either reduced or eliminated, the specific activities of phosphoenolpyruvate carboxylase, malate dehydrogenase, fumarase, fumarate reductase, and NADH:fumarate oxidoreductase were increased substantially, and the acid products were succinate, acetate, and formate. Organisms grown under the latter conditions also contained increased quantities of b- and c-type cytochromes, some of which were low-potential cytochromes. These low-potential cytochromes were reduced by NADH and oxidized by fumarate, and hence, appeared to be components of NADH:fumarate oxidoreductase. Our results indicate that in H. parasuis, growing aerobically in medium containing glucose, the sole function of the reductive pathway is to provide intermediates for biosynthetic processes, and oxygen is the preferred electron acceptor. As the supply of oxygen is reduced or eliminated, the reductive pathway becomes more involved in NAD+ recycling and fumarate becomes the acceptor. In effect, irrespective of the oxygen supply, the growth of H. parasuis is absolutely dependent upon the presence of an electron transport system. Key words: Haemophilus parasuis, cytochromes, enzymes, acids.

scholarly journals Tricarboxylic acid-cycle and related enzymes in restricted facultative methylotrophs

1975 ◽  
Vol 148 (3) ◽  
pp. 505-511 ◽  
Author(s):  
J Colby ◽  
L J Zatman
Keyword(s):  
Succinate Dehydrogenase ◽  
Dehydrogenase Activity ◽  
Isocitrate Dehydrogenase ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Methylotrophic Bacteria ◽  
Acid Cycle ◽  
Oxoglutarate Dehydrogenase ◽  
Specific Activities ◽  
Succinyl Coa Synthetase

The isolation is described of pure cultures of three non-methane-utilizing methylotrophic bacteria which, together with the previously described Bacillus PM6, have a very limited range of growth substrates; these organisms are designated “restricted facultative’ methylotrophs. Two of these isolates, W6A and W3A1, grow only on glucose out of 50 non-C1 compounds tested, whereas the third isolate S2A1 and Bacillus PM6 grow on betaine, glucose, gluconate, alanine, glutamate, citrate and nutrient agar, but not on any of a further 56 non-C1 compounds. Crude sonic extracts of trimethylamine-grown and glucose-grown W6A and W3A1 isolates, and of trimethylamine-grown C2A1 (an obligate methylotroph) contain (i) no detectable 2-oxogltarate dehydrogenase activity, (ii) very low or zero specific activities of succinate dehydrogenase and succinyl-CoA synthetase and (iii) NAD+-dependent isocitrate dehydrogenase activity. Extracts of trimethylamine-grown PM6 and S2A1 methylotrophs have (i) very low 2-oxoglutarate dehydrogenase specific activities, (ii) comparatively high specific activities of succinate dehydrogenase, malate dehydrogenase and succinyl-CoA synthetase and (iii) NADP+-dependent isocitrate dehydrogenase activity but no NAD+-dependent isocitrate dehydrogenase activity. The activities of most of these enzymes are increased during growth on glucose, alanine, glutamate or citrate, but only very low 2-oxoglutarate dehydrogenase activities are present under all growth conditions. The restricted facultative methylotrophs grow on certain non-C1 compounds in the absence of 2-oxoglutarate dehydrogenase and, in some cases, of other enzymes of the tricarboxylic acid cycle; these lesions cannot therefore be the sole cause of obligate methylotrophy.

Use of 14CO2 in estimating rates of hepatic gluconeogenesis

1992 ◽  
Vol 263 (1) ◽  
pp. E36-E41 ◽  
Author(s):  
E. Esenmo ◽  
V. Chandramouli ◽  
W. C. Schumann ◽  
K. Kumaran ◽  
J. Wahren ◽  
...  
Keyword(s):  
Blood Glucose ◽  
Specific Activity ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Hepatic Gluconeogenesis ◽  
Acid Cycle ◽  
Specific Activities ◽  
Glucose Formation

Estimating the rate of hepatic gluconeogenesis in vivo from the incorporation of 14C from 14CO2 into glucose requires determination of the rates in liver of equilibration of oxaloacetate with fumarate, conversion of oxaloacetate to phosphoenolpyruvate (PEP), and conversion of PEP to pyruvate, all relative to the rate of tricarboxylic acid cycle flux. With the use of a model of mitochondrial metabolism and gluconeogenesis, expressions are derived relating specific activity of carboxyl of PEP from 14CO2 to those rates and specific activity of mitochondrial CO2. If those rates and specific activity of mitochondrial CO2 are known, specific activity of PEP, calculated using the expressions, should, on a mole basis, be one-half the specific activity of the glucose formed. At steady state, in the 60-h fasted individual, where glucose formation is solely by gluconeogenesis, twice estimated specific activity of PEP should then approximate that of blood glucose. Estimates of relative rates in 60-h fasted humans, previously made from distribution of 14C in glutamate from phenylacetylglutamine excreted when [3-14C]lactate and phenylacetate were given, were applied to the expressions. Specific activity of mitochondrial CO2 was equated to that of CO2 expired by 60-h fasted subjects given NaH14CO3 and alpha-[1-14C]ketoisocaproate. Predicted specific activities approximated actual specific activities of blood glucose when NaH14CO3 was administered. alpha-[1-14C]ketoisocaproate administrations gave underestimates. This is attributable to differences between specific activities of hepatic mitochondrial CO2 and expired CO2, which is evidenced by higher incorporations of 14C in glucose than in expired CO2 from alpha-[1-14C]ketoisocaproate than from NaH14CO3.(ABSTRACT TRUNCATED AT 250 WORDS

PARTICIPATION OF THE GLYOXYLATE CYCLE IN THE METABOLISM OF ETHANOL BY CASTOR BEAN ENDOSPERM TISSUES

1966 ◽  
Vol 44 (4) ◽  
pp. 423-432 ◽  
Author(s):  
Carol A. Peterson ◽  
E. A. Cossins
Keyword(s):  
Castor Bean ◽  
Tricarboxylic Acid Cycle ◽  
Glyoxylate Cycle ◽  
Experimental Period ◽  
Tricarboxylic Acid ◽  
Ethanol Metabolism ◽  
Short Term ◽  
Tissue Slices ◽  
Acidic Amino Acids ◽  
The Glyoxylate Cycle

The kinetics and pathway of ethanol metabolism in endosperm tissues of the germinating castor bean-have been studied by incubating tissue slices with micromolar quantities of ethanol-1-14C and ethanol-2-14C. In short term experiments, ethanol-14C was incorporated into the organic acids and acidic amino acids. When the experimental period was increased up to 1 hour, large amounts of ethanol-2-14C were incorporated into the sugars, and ethanol-1-14C was extensively incorporated into the respiratory carbon dioxide. Incorporation of ethanol-14C was stimulated by incubation of the tissues with glyoxylate. Ethanol metabolism was markedly inhibited by iodoacetate and malonate. These inhibitors also changed the distribution of14C in the products isolated. Isotopic competition studies indicated that ethanol was incorporated into the acids of the glyoxylate and the tricarboxylic acid cycles at rates substantially lower than acetate.The results are interpreted as being consistent with a metabolism of ethanol mainly via the glyoxylate cycle with some cycling of ethanol carbon through the tricarboxylic acid cycle.

In vitro metabolism by turtle heart mitochondria

1964 ◽  
Vol 206 (5) ◽  
pp. 980-984 ◽  
Author(s):  
H. J. Mersmann ◽  
C. A. Privitera
Keyword(s):  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Chrysemys Picta ◽  
High Rate ◽  
Heart Mitochondria ◽  
Short Term ◽  
Acid Cycle ◽  
Atp Level ◽  
Turtle Heart

Both quantitative and qualitative aspects of the metabolic nature of heart mitochondria in the turtle, Chrysemys picta, have been investigated. Optimal in vitro incubation conditions for oxidative phosphorylation were approximated by alteration of the concentration of various components while using key substrates. The following has been established: exogenous cytochrome c is a necessary component; with an incubation volume of 2 ml the 2-µmole level of F– is the most suitable; 40 µmoles Pi is requisite for substrates metabolized at a high rate; 10 µmoles Mg++ is preferred with an initial ATP level of 3.2 µmoles. Succinate and malate were metabolized at similar rates (18 µmoles/mg N hr). Isocitrate was metabolized at about one-half this rate, whereas addition of NAD increased isocitrate oxidation to the succinate rate. α-Ketoglutarate was utilized at nearly 2.5 times the rate of succinate. Pyruvate oxidation (roughly one-third the succinate rate) increased about ninefold upon addition of 1 µmole malate. ß-Hydroxybutyrate was metabolized at the same rate as succinate. Addition of 1 µmole malate increased this rate nearly fivefold. Citrate apparently was not metabolized. Theoretical P/O ratios were approached with all metabolized substrates. Short-term starvation caused a decrease in both oxidation and phosphorylation. The capacity of turtle heart mitochondria to metabolize intermediates of the tricarboxylic acid cycle appears to be equivalent to that of mitochondria of mammalian cardiac muscle heretofore published.

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