In Vitro Lactate Metabolism by Ruminal Ingesta

Ruminal ingesta (300 ml) obtained from a fistulated cow fed alfalfa hay (H), 3.6 kg of grain mixture with corn silage fed ad libitum (S), 2.5:1 grain-alfalfa hay mixture (G), or a 2.5:1 grain-alfalfa hay mixture providing 545 g of sodium and calcium lactate daily (L) were incubated for 8 hr with nonpolymerized sodium lactate or 17% polymerized lactic acid neutralized to pH 6.7. Polymerization had no effect on the rate of lactate utilization. The initial rates of lactate metabolism for the H, G, S, and L ingesta were 0.72, 0.95, 1.8, and 3.4 meq per 100 ml of rumen fluid per hr, respectively. Lactate-2-(14)C was incubated for 4 hr with each type of ruminal ingesta. Of the label recovered in the volatile fatty acids (VFA), 74.1, 61.2, 49.3, and 38.9% was recovered in acetate, and 9.4, 19.8, 23.3, and 51.9% was recovered in propionate with H, G, S, and L ingesta, respectively. The balance of label was distributed between butyrate and valerate. The titratable VFA did not follow this pattern of production. With the hay ingesta, lactate metabolism resulted in a net loss of acetate and a large increase in butyrate. Little propionate was produced. The G, S, and L ingesta metabolized lactate to yield progressively more propionate and less butyrate. Evidence was gathered to suggest that acetate was the primary end product of lactate metabolism but that oxidation of lactate to pyruvate dictated the synthesis of butyrate from acetate to maintain an oxidation-reduction balance. It was noted that acetate and butyrate production from lactate was pH-dependent, with acetate production maximal at pH 7.4 and butyrate at 6.2. Propionate production was largely unaffected within this pH range.     

Effect of L-methionine and S-adenosylmethionine on growth of an adenine mutant of Saccharomyces cerevisiae

A pink, adenine-requiring yeast utilized adenine, hypoxanthine, or S-adenosylmethionine (SAM), in quantities up to 3 mumoles per 100 ml of medium, as equivalent sources of purine for cell growth, but not methylthioadenosine or S-adenosylhomocysteine. Utilization of SAM for growth was inhibited by the presence of l-methionine in quantities greater than 0.6 mumole per 100 ml of medium. However, 6 mumoles of l-methionine had no effect on growth when adenine or hypoxanthine was the source of purine. These sources also reversed the inhibitory effects of 6 mumoles of the amino acid on the utilization of SAM. The presence of 400 mumoles of the amino acid resulted in some inhibition of growth when the organisms were grown with adenine, hypoxanthine, or adenine plus SAM but had no effect on the total uptake of adenine-8-(14)C. Studies on the uptake of radioactivity from a mixture of SAM-adenine-8-(14)C and (3)H-labeled SAM-methyl indicated that these components were taken into the cells at different rates which were altered by the presence of l-methionine. The fixation of (35)S from (35)S-labeled adenosylmethionine into the cells was inhibited by the presence of the amino acid. The cells synthesized and accumulated SAM in the presence of 400 mumoles of l-methionine plus adenine even when exogenous SAM was supplied. Approximately 47% of radioactivity fixed from exogenous SAM-adenine-8-(14)C and 12% from (3)H-labeled SAM-methyl were found in reisolated SAM.     

Metabolism of propionate by sheep liver. Interrelations of propionate and glutamate in aged mitochondria

1. The rate of metabolism of propionate by aged sheep-liver mitochondria in the presence of oxygen + carbon dioxide (95:5) was 5.0 (+/- s.e.m. 0.8) mumoles/mg. of mitochondrial N/hr. 2. When aged in the presence of the mitochondrial supernatant the rate was increased. Mitochondria from 0.33g. of liver, when combined with the corresponding mitochondrial supernatant from 0.08g. of liver, metabolized propionate at a rate of 11.4 (+/- s.e.m. 1.2) mumoles/mg. of mitochondrial N/hr. This rate is comparable with rates previously obtained with aged nuclear-free homogenates. 3. Two factors in the mitochondrial supernatant were detected, which when combined reproduced the effect of the fresh supernatant and prevented loss of activity on aging. One of these was non-diffusible and was recovered by fractionation of the dialysed mitochondrial supernatant with ammonium sulphate. The second factor was present in an ultrafiltrate of fresh mitochondrial supernatant and in boiled mitochondrial supernatant; it was isolated and identified as l(+)-glutamate. 4. The effect of the non-diffusible factor was due to protection of the mitochondria from the aging process, whereas glutamate served both in this capacity and as a direct stimulant of propionate metabolism at low concentration.     

The role of carbon dioxide in the metabolism of adult Haemonchus contortus, in vitro

Adult Haemonchus contortus worms simultaneously excrete and fix CO2. Their initial content of CO2 was measured as 4.55 mumoles/100 mg wet weight and their excretion rate in air as 1 mumol/100 mg wet weight/h for at least 4 h. When the worms were incubated either aerobically or anaerobically with 14CO2 most of the metabolized radioactivity was associated with propan-1-ol and propionate but small amounts were found in succinate and lactate. No radioactivity was associated with ethanol or acetate, two major catabolites of glucose. Stepwise degradation of the metabolized radioactive propanol and propionate showed that all the radioactivity in both compounds was associated with carbon atom no. 1. These results show that H. contortus has much in common with the anaerobic energy metabolism of Ascaris lumbricoides but they are not inconsistent with the utilization of the tricarboxylic acid cycle by the worm. H. contortus worms were found to metabolize their excretory products. When they were incubated with either [2,3-14C]succinate or [2-14C]acetate, 14CO2 was excreted under aerobic but not under anaerobic conditions. These results are consistent with a pathway similar to that used by Ascaris operating aloneunder anaerobic conditions and together with the tricarboxylic acid cycle under aerobic conditions.     

Microbial Assimilation of Hydrocarbons: Phospholipid Metabolism

An analysis of the turnover of the major phospholipids of Micrococcus cerificans growing or nongrowing cultures. The turnover rates of (14)C-PE and (14)C-PE were 61.5% of the total phospholipid, exhibited no significant rate of turnover in either growing or nongrowing cultures. The turnover rates of PE-(14)C and PE-(32)P were 3.2% per hr and 1.2% per hr, respectively. Phosphatidylglycerol (PG) exhibited a turnover rate of 11% and 7.7% per hr for (14)C and (32)P, respectively, indicating an extremely slow metabolism. PG metabolism was examined in greater detail, and the data indicated a preferential 75% incorporation of glycerol-1,3-(14)C into the unacylated portion of the PG molecule. The turnover of cardiolipin (CL) was extremely slow in growing cells whereas nongrowing cells exhibited a 30% and 36% increase per hr for (14)C-Cl and (14)C-CL, respectively. Glycerol-1,3-(14)C was not converted to phospholipid fatty acid carbon; all radioactivity appeared only in the water-soluble backbone of the phospholipids. The kinetics of assimilation of hexadecane-1-(14)C into cellular lipids is presented. Radioactivity in neutral lipid increased approximately sevenfold over the growth cycle, whereas radioactivity in phospholipid increased 50-fold during the same time period. The incorporation of radioactive fatty acids derived from the direct oxidation of hexadecane-1-(14)C demonstrated differential kinetics of assimilation into PE, PG, and CL. The results indicated a rapid turnover of phospholipid fatty acids in M. cerificans growing at the expense of hexadecane.     

Oxygen uptake and lactate production by Schistosoma mansoni cercaria, cercarial body and tail, and schistosomule.

1. Oxygen consumption by Schistosoma mansoni cercarial bodies varies, with the batch of organisms, the incubation media and the temperature (27-37 degrees C), from 27.4 +/- 3.4 to 55.0 +/- 4.8 microliters O2/mg larval protein per hr. It is proportional to the concentration of organisms incubated, up to 25,000/ml, as calculated from whole protein. 2. Oxygen uptake by cercariae is inhibited by 5.6 mM glucose in the incubation media, a concentration that stimulates the respiration of cercarial bodies. 3. No significant differences in the oxygen uptake were presented by cercarial bodies with and without glycocalyx or glandular secretions, or devoid of all of them. 4. Inhibitors of the Krebs cycle and the respiratory chain, and uncoupling agents influence the oxygen uptake by cercariae, cercarial bodies and schistosomules to the same extent. 5. The permeability change presented by transformed larvae had no influence on the excretion of lactate by cercarial bodies, which is about 0.3 mumoles/mg protein per hr and remains constant for 5 hr; under nitrogen, this amount increased 70%. Cercariae in anaerobiosis, however, excreted as much as 15 times more lactate than under air. 6. Lactic dehydrogenases of cercariae, cercarial bodies and tails, and schistosomules are of the muscle type and do not change during the transformation.     

Metabolic engineering of a non-allosteric citrate synthase in an Escherichia coli citrate synthase mutant

This study examined the organization of the Krebs tricarboxylic acid (TCA) cycle by metabolic engineering and high-resolution ¹³C NMR. The oxidation of [1,2,3-¹³C]propionate to glutamate via the TCA cycle was measured in wild-type (WT) and a citrate synthase mutant (CS^?) strain of Escherichia coli transformed with allosteric E. coli citrate synthase (ECCS) or non-allosteric pig citrate synthase (PCS). The ¹³C fractional enrichment in glutamate C-2, C-3, and C-4 in ECCS and PCS were similar; although quantitative differences in total citrate synthase activity and total C-4 labeling of glutamate were observed in ECCS and PCS. Allosteric ECCS cells contained 10-fold less total enzyme activity than PCS but only 50% less total labeling in glutamate C-4 and equivalent doubling times. The observed spectra were mathematically fitted using an iterative procedure(TCACALC) and yielded an acetate/succinyl-CoA flux ratio of 10 for both ECCS and PCS, a result that is in agreement with the isotopomer analyses of the ¹³C spectra of cells presented with [3-¹³C] propionate or [2-¹³C]propionate. The results are consistent with the presence of an allosteric citrate synthase in ECCS and a non-allosteric citrate synthase in PCS. The former maintains TCA cycle flux via alternative propionate pathways activated by positive allosteric mechanisms and the latter via elevated enzyme levels.     

Metabolism of Propane, n-Propylamine, and Propionate by Hydrocarbon-Utilizing Bacteria

Studies were conducted on the oxidation and assimilation of various three-carbon compounds by a gram-positive rod isolated from soil and designated strain R-22. This organism can utilize propane, propionate, or n-propylamine as sole source of carbon and energy. Respiration rates, enzyme assays, and ¹⁴CO₂ incorporation experiments suggest that propane is metabolized via methyl ketone formation; propionate and n-propylamine are metabolized via the methylmalonyl-succinate pathway. Isocitrate lyase activity was found in cells grown on acetate and was not present in cells grown on propionate or n-propylamine. ¹⁴CO₂ was incorporated into pyruvate when propionate and n-propylamine were oxidized in the presence of NaAsO₂, but insignificant radioactivity was found in pyruvate produced during the oxidation of propane and acetone. The n-propylamine dissimilatory mechanism was inducible in strain R-22, and amine dehydrogenase activity was detected in cells grown on n-propylamine. Radiorespirometer and ¹⁴CO₂ incorporation studies with several propane-utilizing organisms indicate that the methylmalonyl-succinate pathway is the predominant one for the metabolism of propionate.     

Metabolism of [3-13C]pyruvate in TCA cycle mutants of yeast.

The utilization of pyruvate and acetate by Saccharomyces cerevisiae was examined using 13C and 1H NMR methodology in intact wild-type yeast cells and mutant yeast cells lacking Krebs tricarboxylic acid (TCA) cycle enzymes. These mutant cells lacked either mitochondrial (NAD) isocitrate dehydrogenase (NAD-ICDH1),alpha-ketoglutarate dehydrogenase complex (alpha KGDC), or mitochondrial malate dehydrogenase (MDH1). These mutant strains have the common phenotype of being unable to grow on acetate. [3-13C]-Pyruvate was utilized efficiently by wild-type yeast with the major intermediates being [13C]glutamate, [13C]acetate, and [13C]alanine. Deletion of any one of these Krebs TCA cycle enzymes changed the metabolic pattern such that the major synthetic product was [13C]galactose instead of [13C]glutamate, with some formation of [13C]acetate and [13C]alanine. The fact that glutamate formation did not occur readily in these mutants despite the metabolic capacity to synthesize glutamate from pyruvate is difficult to explain. We discuss the possibility that these data support the metabolon hypothesis of Krebs TCA cycle enzyme organization.     

Partial characterization of the pathway from propionate to acetate in the cabbage looper,Trichoplusia ni

Experiments were performed to characterize the metabolism of propionate to acetate in the cabbage looper Trichoplusia ni and correlate the results with vitamin B₁₂ levels. Fourth and fifth instar larvae contain 2–4 pg vitamin B₁₂/mg dry wt whereas pupae and adults do not contain detectable amounts. In vivo studies as a function of time in larvae, pupae and adults gave evidence that [2-¹⁴C]propionate was converted to 3-hydroxypropionate and then to acetate, which subsequently labeled Krebs cycle intermediates. Radioactivity from [1-¹⁴C]propionate was recovered only in the propionate and 3-hydroxypropionate fractions, and not in acetate or Krebs cycle intermediates, suggesting that carbon 1 of propionate was lost as carbon dioxide and that carbons 2 and 3 of propionate were retained during conversion to acetate. The enzymes of this pathway were located entirely in the mitochondrial fraction. Cyanide inhibited the metabolism of propionate to 3-hydroxypropionate and acetate in mitochondrial preparations, whereas carbon monoxide did not. [2,3-¹⁴C]Acrylic acid was metabolized to 3-hydroxypropionate, which is consistent with a dehydrogenase converting propionate to acrylate which is then hydrated to 3-hydroxypropionate and then oxidized and decarboxylated to acetate.     

Carbon-13 Nuclear Magnetic Resonance Study of Metabolism of Propionate by Escherichia coli

We have evaluated the use of [1,2-13C2]propionate for the analysis of propionic acid metabolism, based on the ability to distinguish between the methylcitrate and methylmalonate pathways. Studies using propionate-adapted Escherichia coli MG1655 cells were performed. Preservation of the 13C-13C-12C carbon skeleton in labeled alanine and alanine-containing peptides involved in cell wall recycling is indicative of the direct formation of pyruvate from propionate via the methylcitrate cycle, the enzymes of which have recently been demonstrated in E. coli. Additionally, formation of 13C-labeled formate from pyruvate by the action of pyruvate-formate lyase is also consistent with the labeling of pyruvate C-1. Carboxylation of the labeled pyruvate leads to formation of [1,2-13C2]oxaloacetate and to multiply labeled glutamate and succinate isotopomers, also consistent with the flux through the methylcitrate pathway, followed by the tricarboxylic acid (TCA) cycle. Additional labeling of TCA intermediates arises due to the formation of [1-13C]acetyl coenzyme A from the labeled pyruvate, formed via pyruvate-formate lyase. Labeling patterns in trehalose and glycine are also interpreted in terms of the above pathways. The information derived from the [1, 2-13C2]propionate label is contrasted with information which can be derived from singly or triply labeled propionate and shown to be more useful for distinguishing the different propionate utilization pathways via nuclear magnetic resonance analysis.     

Kinetics of Acetate Metabolism during Sludge Digestion

The quantitative contribution of acetic acid to methane production was studied by use of C(14)-labeled acetic acid. Samples of domestic sewage sludge were incubated anaerobically in Warburg vessels. The rate of methane production in the vessels was 0.033 mumoles per ml per min. C(14)-labeled acetic acid was added, and the turnover rate was calculated. The pool size of acetic acid in the sludge was 4.7 mumoles/ml. The turnover rate was 0.0052 min(-1), giving a rate of formation of acetic acid of 0.024 mumoles per ml per min. Under these conditions, acetic acid would account for approximately 73% of the methane produced by the sludge. Acetic acid was found to exist primarily in an extracellular pool. The turnover rate of the extracellular pool was rapid, and it was concluded that most of the acetic acid must be metabolized to methane by a specialized microflora not involved in the formation of acetic acid.     

Metabolism of D-fructose by Arthrobacter pyridinolis

Previous studies showed that Arthrobacter pyridinolis can transport and utilize d-glucose only after prior growth on certain Krebs cycle intermediates. In contrast, we found that d-fructose was taken up and metabolized by A. pyridinolis without special prior conditions of growth. d-Fructose was first converted to d-fructose-1-phosphate by a phosphoenolpyruvate (PEP):D-fructose phosphotransferase. This activity required both supernatant and pellet fractions from d-fructose-grown cells centrifuged at 150,000 x g. The d-fructose-1-phosphate formed was converted to d-fructose-1, 6-diphosphate. Mutants deficient in PEP:d-fructose phosphotransferase and d-fructose-1-phosphate kinase, or d-fructose-1, 6-diphosphatase (FDPase) were unable to grow on d-fructose but retained the normal ability to use d-glucose. Mutants forming reduced amounts of FDPase were completely unable to grow on d-fructose but were still capable of limited growth on Krebs cycle intermediates. A requirement for higher levels of FDPase for growth on d-fructose than for growth on Krebs cycle intermediates was also indicated by the higher specific activities of FDPase in d-fructose-grown cells than in cells grown on l-malate or amino acids.     

Glucose Utilization by Euglena gracilis var. bacillaris at Higher pH*

SYNOPSIS. Euglena gracilis var. bacillaris is able to grow luxuriantly on glucose in a mineral salts medium at pH 6.8–7.1 following an adaptation period of about 200 hr. If adapted cells are used as an inoculum or if 0.1% glycine is included in the medium, the lag is shortened to 70–100 hr. Inclusion of 0.1% acetate in the medium produces a diphasic growth pattern, with acetate being metabolized first, followed by the later (about 400 hr) utilization of the glucose. Glucose utilization was found to be sensitive to pH as compared to growth on ethyl alcohol. However, glycine partially overcame this sensitivity. Glycine is maximally stimulatory with regard to growth on glucose at pH 7.0 at a concentration of 0.03%, thus suggesting that it functions as a sparking substance. Glycine markedly stimulates the assimilation of ¹⁴C-glucose. A number of Krebs cycle acids and amino acids were also found to stimulate ¹⁴C-glucose assimilation at neutral pH. Adaptation to glucose utilization at neutral pH was due to the appearance of mutants able to grow more rapidly under these conditions. The nature of this mutation was not determined.     

In Vivo and In Vitro Studies on gamma-Aminobutyric Acid Metabolism with the Radish Plant (Raphanus sativus, L.)

Labeled glutamate was rapidly converted to γ-aminobutyrate in intact, excised radish (Raphanus sativus L., var. Champion) leaves. Labeled γ-aminobutyrate was metabolized via succinate and the Krebs cycle and was not carboxylated to form glutamate. Administration of carbon-14 and tritium-labeled succinate indicated that less than 10% of the γ-aminobutyrate formation occurs by amination of succinic semialdehyde. Therefore, most γ-aminobutyrate formation must be via glutamate decarboxylation.     

Metabolism of propionate by sheep liver. Oxidation of propionate by homogenates

1. The rate and stability to aging of the metabolism of propionate by sheep-liver slices and sucrose homogenates were examined. Aging for up to 20min. at 37° in the absence of added substrate had little effect with slices, whole homogenates or homogenates without the nuclear fraction. 2. Metabolism of propionate by sucrose homogenates was confined to the mitochondrial fraction, but the mitochondrial supernatant (microsomes plus cell sap) stimulated propionate removal. 3. The rate of propionate metabolism by liver slices was higher in a high potassium phosphate–bicarbonate medium [0·88(±s.e.m. 0·16)μmole/mg. of N/hr.] than in Krebs–Ringer bicarbonate medium [0·44(±s.e.m. 0·13)μmole/mg. of N/hr.]. 4. Metabolism of propionate by sucrose homogenates freed from nuclei was dependent on the presence of oxygen, carbon dioxide and ATP. Propionate removal was stimulated 250% by Mg²⁺ ions and 670% by cytochrome c. 5. In the complete medium 2·39(±s.e.m. 0·15)μmoles of propionate were consumed/mg. of N/hr. 6. The ratio of oxygen consumption to propionate utilization was sufficient to account for the complete oxidation of half the propionate consumed. 7. The only products detected under these conditions were succinate, fumarate and malate. Propionate had no effect on the production of lactate from endogenous sources and did not itself give rise to lactate. 8. Methylmalonate did not accumulate when propionate was metabolized and was not oxidized. It was detected as an intermediate in the conversion of propionyl-CoA into succinate. The rate of this reaction sequence was adequate to account for the rate of propionate metabolism by sucrose homogenates or slices, provided that the rate of formation of propionyl-CoA was not limiting. 9. The methylmalonate pathway was predominantly a mitochondrial function. 10. The metabolism of propionate appeared to be dependent on active oxidative phosphorylation.     

Metabolism of propionate to acetate in the cockroach Periplaneta americana

Carbon-13 NMR and radiotracer studies were used to determine the precursor to methylmalonate and to study the metabolism of propionate in the cockroach Periplaneta americana. [3,4,5-13C3]Valine labeled carbons 3, 4, and 26 of 3-methylpentacosane, indicating that valine was metabolized via propionyl-CoA to methylmalonyl-CoA and served as the methyl branch unit precursor. Potassium [2-13C]propionate labeled the odd-numbered carbons of hydrocarbons and potassium [3-13C]propionate labeled the even-numbered carbons of hydrocarbons in this insect. This labeling pattern indicates that propionate is metabolized to acetate, with carbon-2 of propionate becoming the methyl carbon of acetate and carbon-3 of propionate becoming the carboxyl carbon of acetate. In vivo studies in which products were separated by HPLC showed that [2-14C]propionate was readily metabolized to acetate. The radioactivity from sodium [1-14C]propionate was not incorporated into succinate nor into any other tricarboxylic acid cycle intermediate, indicating that propionate was not metabolized via methylmalonate to succinate. Similarly, [1-14C]propionate did not label acetate. An experiment designed to determine the subcellular localization of the enzymes involved in converting propionate to acetate showed that they were located in the mitochondrial fraction. Data from both in vivo and in vitro studies as a function of time indicated that propionate was converted directly to acetate and did not first go through tricarboxylic acid cycle intermediates. These data demonstrate a novel pathway of propionate metabolism in insects.     

Effect of propionate on the utilization of nitrogen from 15NH4Cl for urea synthesis in hepatocytes isolated from sheep liver

1. The effect of ornithine (2.0 mM) and propionate (5.0 mM) on the utilization of N from 15NH4Cl (5.0 mM) for urea synthesis in hepatocytes isolated from sheep liver was investigated. 2. The capacity of sheep hepatocytes to utilize [15N]ammonia in the absence of the other exogenous substrates was very low and amounted 132 +/- 37.3 mumol/hr per 1 g dry wt. 3. Ornithine failed to affect the total [15N]ammonia uptake and total urea synthesis, but at the same time it markedly increased the utilization of [15N]ammonia for ureagenesis and diminished the rate of urea synthesis from endogenous sources. 4. Propionate markedly increased total [15N]ammonia utilization and total urea formation; this increase resulted from the rise of ammonia utilization for urea synthesis and it was similar in the presence or absence of ornithine. 5. The capacity of sheep liver cells to utilize ammonia in the presence of propionate (in the presence or absence of ornithine) amounted to 256 mumol/hr per 1 g dry wt, thus being similar to the values in vivo. 6. It is concluded that in sheep hepatocytes both ornithine and propionate stimulate the utilization of ammonia for urea synthesis and these effects take place independently and occur by different mechanisms.     

Effects of deflazacort and the L-6485 metabolite on epiphyseal cartilage carbohydrate metabolism: comparison with prednisone

Male Sprague-Dawley rats were injected with 1 mg/100 g bw/day of prednisone, 1.25 mg/1--g bw/day of deflazacort, or its metabolite, for a period of 20 days. Epiphyseal cartilage slices were incubated in a modified Krebs Ringer bicarbonate buffer, at 37 degrees C for 60 min, with either 14C-1- or 14C-6-glucose to quantitate both the absolute and relative rates of pentose shunt versus aerobic and anaerobic glycolytic activity, respectively. Measurements of both total and radioactive glucose uptake, lactate production and 14CO2 generation were expressed as either mumoles or DPM/mg cellular DNA/hr, respectively. This study demonstrated: (1) chronic prednisone administration decreased anaerobic glycolysis (glucose uptake and lactate production) 3-fold (P less than 0.01); (2) prednisone on a chronic basis produced no measured alteration in either the pentose shunt or Kreb's cycle activity; (3) both deflazacort and the deflazacort metabolite significantly stimulated (P less than 0.02) anaerobic glycolytic activity in epiphyseal cartilage tissue. In contrast to prednisone, the administration of either deflazacort or its L-6485 metabolite did not inhibit the glycolytic pathway of metabolism so necessary for epiphyseal cartilage growth and mineralization.     

Glucose and Fructose Metabolism in a Phosphoglucoisomeraseless Mutant of Saccharomyces cerevisiae

A mutant of Saccharomyces cerevisiae deficient in phosphoglucoisomerase (EC 5.3.1.9) is described. It does not grow on glucose or sucrose but does grow on galactose or maltose. Addition of glucose to cultures growing on fructose, mannose, or acetate arrests further growth without altering viability; removal of glucose permits resumption of growth. Glucose causes accumulation of nearly 30 mumoles of glucose-6-phosphate per g (wet weight) of cells and suppresses synthesis of ribonucleic acid. Inhibition of growth by glucose does not appear to be due to a loss of adenosine triphosphate or inorganic orthophosphate. The mutant, however, utilizes glucose-6-phosphate produced intracellularly. Release of carbon dioxide from specifically labeled glucose suggests a C-l preferential cleavage. The kinetics of glucose-6-phosphate accumulation during glucose utilization in the mutant is not consistent with the notion that the utilization of glucose is controlled by glucose-6-phosphate.