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PubMed1977

Influence of 1,2,3-benzene-tricarboxylate on pyruvate metabolism in rat-liver mitochondria.

1,2,3-Benzene-tricarboxylate, a known inhibitor of the mitochondrial tricarboxylate carrier, was found to inhibit pyruvate carboxylation as well as the transport of citrate out of the matrix in rat liver mitochondria incubated with pyruvate. The inhibition of pyruvate carboxylation was observed with both intact mitochondria and with the solubilized pyruvate carboxylase. The inhibition of the pyruvate carboxylase by 1,2,3-benzene-tricarboxylase was not mediated via one of the parameters known to regulate the activity of the enzyme and therefore a direct inhibition of the enzyme by the tricarboxylate was assumed. Since the pyruvate carboxylase is exclusively localized in the mitochondrial matrix space it was concluded that 1,2,3-benzene-tricarboxylate penetrates into this compartment.

Acetyl Coenzyme A↗
PubMed1985

Stimulation by 3-hydroxybutyrate of pyruvate carboxylation in mitochondria from rat liver.

Isolated rat liver mitochondria incubated in the presence of 3-hydroxybutyrate display a markedly increased rate of pyruvate carboxylation as measured by malate and citrate production from pyruvate. The stimulation was demonstrable both with exogenously added pyruvate, even at saturating concentration, and with pyruvate intramitochondrially generated from alanine. The concentration of DL-3-hydroxybutyrate required for half-maximal stimulation amounted to about 1.5 mM. The intramitochondrial ATP/ADP ratio as well as the matrix acetyl-CoA level was found to remain unchanged by 3-hydroxybutyrate exposure, which, however, lowered the absolute intramitochondrial contents of the respective adenine nucleotides. The effects of 3-hydroxybutyrate were diminished by the concomitant addition of acetoacetate. Moreover, a direct relationship between mitochondrial reduction by proline and the rate of pyruvate carboxylation was observed. The results seem to indicate that the mitochondrial oxidation--reduction state might be involved in the expression of the 3-hydroxybutyrate effect. As to the physiological relevance of the findings, 3-hydroxybutyrate could be shown to activate pyruvate carboxylation in isolated hepatocytes.

Animals↗
PubMed1988

Control of pyruvate carboxylase activity by the pyridine-nucleotide redox state in mitochondria from rat liver.

Pyruvate carboxylation by isolated mitochondria from rat liver is inhibited by t-butylhydroperoxide in a fully reversible manner. The rate of malate formation at 10 mM pyruvate was decreased by some 80% by 30 microM t-butylhydroperoxide. The effective peroxide concentration was dependent on the mitochondrial hydrogen supply, being increased to about 120 microM in the presence of 50 microM palmitoylcarnitine. Regarding the mechanism(s) of the t-butylhydroperoxide action, pyruvate transport and intramitochondrial energy or activator supply are unlikely involved, because the effect also took place with alanine as the substrate and was not accompanied by a change in the intramitochondrial levels of adenine nucleotides and acetyl-CoA respectively. However, t-butylhydroperoxide caused a rapid fall in the 3-hydroxybutyrate/acetoacetate ratio and a marked increase in the oxidized glutathione content. Therefore, experiments were designed to disclose the participation of the respective redox couples in the expression of pyruvate carboxylase activity. From measurements of NADPH, NADH, oxidized and reduced glutathione contents of mitochondria incubated under a variety of conditions, evidence has been obtained indicating that the mitochondrial NADH supply represents an important factor in the regulation of pyruvate carboxylase activity. The results presented seemingly provide a new basis for the understanding of the functional relationship between beta-oxidation and pyruvate carboxylation.

3-Hydroxybutyric Acid↗
PubMed1990

A radical-chemical route to acetyl-CoA: the anaerobically induced pyruvate formate-lyase system of Escherichia coli.

Anaerobically growing Escherichia coli cells contain the enzyme pyruvate formate-lyase which catalyses the non-oxidative cleavage of pyruvate to acetyl-CoA and formate. The enzyme is subject to interconversion between inactive and active forms. The active form contains an oxygen-sensitive organic free radical located on the polypeptide chain which is essential for catalysis. It affords a novel homolytic C-C bond cleavage of the pyruvate substrate. The radical is generated by an iron-dependent converter enzyme which requires reduced flavodoxin and adenosyl methionine as co-substrates and pyruvate as a positive allosteric effector. A second converter enzyme, also iron-dependent, accomplishes the removal of the radical. This post-translational interconversion cycle controls the activity state of pyruvate formate-lyase in the anaerobic cell. Anaerobic conditions also regulate pyruvate formate-lyase at the level of gene expression. Multiple promoters are responsible for effecting a twelve to fifteen fold induction and they are coordinately controlled in response to the oxygen and metabolic status of the cell by sequences which are located far upstream of the pfl coding region. The transcription factor Fnr has been identified as being responsible for part of the anaerobic control of pfl expression, probably through direct interaction with the upstream sequences. In contrast, the expression of the gene encoding the first iron-dependent converter enzyme is unaffected by anaerobiosis and is independent of the Fnr protein.

Acetyl Coenzyme A↗
PubMed1977

Fermentative metabolism of pyruvate by Rhodospirillum rubrum after anaerobic growth in darkness.

Rhodospirillum rubrum grew anaerobically in darkness and fermented sodium pyruvate by a pyruvate formate-lyase reaction. During 30 min of anaerobic dark or light incubation with sodium pyrivate, crude extracts from fermentatively grown cells produced about 6 micronmol of acetylphosphate and formate per mg of protein in reactions performed at pH 8.3. Cell extracts also catalyzed the exchange of sodium [14C]formate into sodium pyruvate at an apparent pH optimum of 7.3 to 7.5, but only about 2.5 micronmol of acetylphosphate was produced at this lower pH value. R. rubrum may also form pyruvate:ferredoxin oxidoreductase activity, as evidenced by low bicarbonate exchange activity. However, its participation in pyruvate metabolism in anaerobic dark-grown cells was not understood. During anaerobic, dark growth with pyruvate, formate was an intermediate in H2 and CO2 gas evolution. In contrast with H2 production by a light-dependent H2-nitrogenase system in photosynthetically grown cells, H2 formation in fermenting R. rubrum occurred through a carbon monoxide-sensitive formic hydrogenlyase reaction not influenced by light.

Aldehyde Oxidoreductases↗
PubMed1993

Integration host factor is required for anaerobic pyruvate induction of pfl operon expression in Escherichia coli.

The expression of the pyruvate formate-lyase gene (pfl) is induced by anaerobic growth, and this is increased further by growth in pyruvate. Previous work has shown that anaerobic induction is strongly dependent on the activator FNR and partially dependent on a second transcription factor, ArcA, while pyruvate induction only required FNR. Anaerobic and pyruvate regulation both require the presence of a 5' nontranslated regulatory sequence which spans approximately 500 bp of DNA. A mobility shift assay was developed to identify proteins that bind to this regulatory region. Several binding activities were separated by heparin agarose chromatography, and one of these activities was characterized and shown to be integration host factor (IHF). Mobility shift and DNase I footprinting experiments defined a single IHF binding site in the pfl promoter-regulatory region. With pfl-lacZ fusions, it could be shown that introduction of a himD mutation abolished pyruvate-dependent induction of anaerobic expression in vivo. The same result was observed when the pfl IHF binding site was mutated. In addition, the partial anaerobic induction of expression found in an fnr strain was completely blocked in an fnr himD double mutant and in an fnr IHF binding site double mutant. Taken together, these data suggest that IHF is necessary for both pyruvate induction and the anaerobic induction mediated by ArcA.

Acetyltransferases↗
PubMed1990

Improved detection of acid mine water stressed coliform bacteria on media containing catalase and sodium pyruvate.

Pure culture suspensions of two strains of exponential and stationary phase Escherichia coli exhibited significant reductions in catalase activity following exposure to acid mine water (AMW). The exogenous addition of catalase (500-2000 U) or sodium pyruvate (0.05-5%) to a nonselective recovery medium resulted in enhanced detection (12- to 465-fold) of AMW-stressed E. coli as compared with recovery on the medium lacking these supplements, whereas addition of 3,3'-thiodipropionic acid failed to improve recovery. Additional in vitro experiments utilizing selective M-FC, mT7, and M-Endo media containing 1000 U catalase or 1.0% pyruvate similarly resulted in improved detection of AMW-stressed cells, with the exception of M-Endo containing pyruvate. Appropriately modified media were then used to analyze an AMW-impacted stream by the membrane filtration technique. Addition of catalase, pyruvate, or a combination of both significantly improved recovery of fecal and total coliforms without promoting growth of noncoliforms. Supplementation of plate count agar with pyruvate and (or) catalase enhanced detection of total heterotrophs. These findings suggest that addition of catalase or pyruvate to standard recovery media may improve detection of coliform and total heterotrophic bacteria in AMW-impacted waters.

Bacterial Proteins↗
PubMed1996

Intracellular lactate- and pyruvate-interconversion rates are increased in muscle tissue of non-insulin-dependent diabetic individuals.

The contribution of muscle tissues of non-insulin-dependent diabetes mellitus (NIDDM) patients to blood lactate appearance remains undefined. To gain insight on intracellular pyruvate/lactate metabolism, the postabsorptive forearm metabolism of glucose, lactate, FFA, and ketone bodies (KB) was assessed in seven obese non-insulin-dependent diabetic patients (BMI = 28.0 +/- 0.5 kg/m2) and seven control individuals (BMI = 24.8 +/- 0.5 kg/m2) by using arteriovenous balance across forearm tissues along with continuous infusion of [3-13C1]-lactate and indirect calorimetry. Fasting plasma concentrations of glucose (10.0 +/- 0.3 vs. 4.7 +/- 0.2 mmol/liter), insulin (68 +/- 5 vs. 43 +/- 6 pmol/liter), FFA (0.57 +/- 0.02 vs. 0.51 +/- 0.02 mmol/liter), and blood levels of lactate (1.05 +/- 0.04 vs. 0.60 +/- 0.06 mmol/liter), and KB (0.48 +/- 0.04 vs. 0.29 +/- 0.02 mmol/liter) were higher in NIDDM patients (P < 0.01). Forearm glucose uptake was similar in the two groups (10.3 +/- 1.4 vs. 9.6 +/ 1.1 micromol/min/liter of forearm tissue), while KB uptake was twice as much in NIDDM patients as compared to control subjects. Lactate balance was only slightly increased in NIDDM patients (5.6 +/- 1.4 vs. 3.3 +/- 1.0 micromol/min/liter; P = NS). A two-compartment model of lactate and pyruvate kinetics in the forearm tissue was used to dissect out the rates of lactate to pyruvate and pyruvate to lactate interconversions. In spite of minor differences in the lactate balance, a fourfold increase in both lactate- (44.8 +/- 9.0 vs. 12.6 +/- 4.6 micromol/min/liter) and pyruvate-(50.4 +/- 9.8 vs. 16.0 +/- 5.0 micromol/min/liter) interconversion rates (both P < 0.01) were found. Whole body lactate turnover, assessed by using the classic isotope dilution principle, was higher in NIDDM individuals (46 +/- 9 vs. 21 +/- 3 micromol/min/kg; P < 0.01). Insights into the physiological meaning of this parameter were obtained by using a whole body noncompartmental model of lactate/pyruvate kinetics which provides a lower and upper bound for total lactate and pyruvate turnover (NIDDM = 46 +/- 9 vs. 108 +/- 31; controls = 21 +/- 3 - 50 +/-13 micromol/min/kg). In conclusion, in the postabsorptive state, despite a trivial lactate release by muscle, lactate- and pyruvate-interconversion rates are greatly enhanced in NIDDM patients, possibly due to concomitant impairment in the oxidative pathway of glucose metabolism. This finding strongly suggest a major disturbance in intracellular lactate/pyruvate metabolism in NIDDM.

Adult↗
PubMedDate not supplied

[Isolation and characterization of a membrane-bound pyruvate dehydrogenase complex from the phototrophic bacterium Rhodospirillum rubrum (author's transl)].

The pyruvate dehydrogenase complex from the photosynthetic bacterium Rhodospirillum rubrum was associated with the membrane fraction both in heterotrophically and photosynthetically grown cells. The complex was separated from the membranes and partially purified by precipitation with MgSO4 and gelfiltration through Sepharose 4B. The purified complex had a specific activity of 1.5-2mumol/min-mg protein and contained the following partial activities: pyruvate dehydrogenase (EC 1.2.4.1), dihydrolipoamide transacetylase (EC 2.3.1.12) and dihydrolipoamide dehydrogenase (EC 1.6.4.3). Contrary to other bacterial pyruvate dehydrogenase complexes, the enzyme complex from R. rubrum revealed no cooperatively between pyruvate binding sites. The kinetic constants (Km) for the overall reaction were (in mM): 0.14 (pyruvate), 0.07 (NAD) and 0.025 (coenzyme A). The Km for thiamine pyrophosphate was dependent on the nature and the concentration of the divalent metal ion (Mn or Mg) present in the reaction mixture, the values ranging from 0.5 to 3 micrometer. NADH was a potent inhibitor (Ki=5 micrometer) of the enzyme complex and the dihydrolipo amide dehydrogenase. The inhibition was competitive with respect to NAD. In addition to its rapid inhibitory effect, NADH also inactivated the enzyme. Cysteine partially protected the enzyme complex against NADH-inactivation. Acetyl-coenzyme A also inhibited the overall reaction (Ki=40 micrometer). The inhibition was dependent on the concentration of coenzyme A, but independent of the concentration of pyruvate. Sugar phosphates, phosphoenolpyruvate, citric acid cycle intermediates and nucleosidephosphates (1 mM) had no pronounced effect on the overall reaction.

Acetyl Coenzyme A↗
PubMed1987

The human pyruvate dehydrogenase complex. Isolation of cDNA clones for the E1 alpha subunit, sequence analysis, and characterization of the mRNA.

cDNA clones corresponding to the entire length of mRNA for the alpha subunit of human pyruvate dehydrogenase (EC 1.2.4.1), the E1 component of the pyruvate dehydrogenase complex, have been isolated from liver cDNA libraries. Two classes of cDNA clones were obtained and these correspond to two forms of pyruvate dehydrogenase E1 alpha mRNA. Both mRNA species have been demonstrated in a variety of human tissues and cultured fibroblasts. The cDNA sequence has been determined and, from it, the protein sequence of the human E1 alpha subunit was deduced. The protein is synthesized with a typical mitochondrial import leader sequence and the peptide bond at which this sequence is cleaved after transport into the mitochondrion has been determined by direct amino acid sequencing of the mature E1 alpha subunit. The human pyruvate dehydrogenase E1 alpha subunit contains identical phosphorylation sites to those found in the corresponding porcine protein. Preliminary studies of pyruvate dehydrogenase E1 alpha mRNA in cultured fibroblasts from patients with severe pyruvate dehydrogenase deficiency have revealed considerable heterogeneity as would be expected from protein studies.

Amino Acid Sequence↗
PubMed1988

In vivo studies of cysteine metabolism. Use of D-cysteinesulfinate, a novel cysteinesulfinate decarboxylase inhibitor, to probe taurine and pyruvate synthesis.

Although several pathways contribute to the catabolism of L-cysteine, the products formed are few--taurine + CO2 and pyruvate + ammonia + sulfate. L-Cysteinesulfinate is a key intermediate that is either decarboxylated to ultimately yield taurine or transaminated to yield pyruvate. There is strong evidence that pyruvate is also formed by several cysteinesulfinate-independent pathways collectively referred to as "cysteine desulfhydrase." The quantitative importance of cysteinesulfinate-independent pathways of taurine synthesis is less clear, but it has been suggested that taurine synthesis from the cysteamine released during phosphopantetheine and CoASH turnover accounts for the high taurine content of tissues with very low levels of cysteinesulfinate decarboxylase activity (e.g. skeletal muscle and heart). In the present studies, the metabolic flux through each of these pathways was quantitated in vivo by monitoring the formation of respiratory 14CO2 in mice administered L-[1-14C]- or L-[3-14C]cyst(e)ine. Mice given 0.05 mmol/kg of L-cystine or 0.5 or 2.5 mmol/kg of L-cysteine catabolize 35, 51, and 72% of the dose, respectively, in 6 h; the relative contribution of taurine synthesis to total catabolism decreases from 63 to 51 to 42% as the L-cyst(e)ine dose is increased. To evaluate the role of L-cysteinesulfinate in taurine synthesis, D-cysteinesulfinate was characterized and used as a metabolism-resistant, potent, and specific inhibitor of cysteinesulfinate decarboxylase. Studies with L-[1-14C]- and L-[3-14C]cysteine in the presence of inhibitor indicate that 85-93% of taurine synthesis occurs from L-cysteinesulfinate: the calculated contribution of the phosphopantetheine pathway is small and may approximate zero. L-Cysteinesulfinate transmamination accounts for 25% of pyruvate synthesis from L-[14C]cystine (0.05 mmol/kg) but only 11% of pyruvate synthesis from L-[14C]cysteine (2.5 mmol/kg). Cysteine desulfhydrase reactions account for most of the pyruvate synthesis.

Animals↗
PubMed1988

Metabolic effects of partial reversal of pyruvate dehydrogenase activity by dichloroacetate in sepsis.

The metabolic effects of dichloroacetate on carbohydrate metabolism were investigated in normal fed, sterile inflammatory, and chronic septic animals. Chronic sepsis, but not sterile inflammation, was associated with elevated plasma, liver, and skeletal muscle lactate concentrations. Sodium dichloroacetate significantly reduced both plasma and intracellular pyruvate and lactate concentrations in all conditions examined, while plasma glucose concentrations remained unchanged. Decreased tissue metabolite concentrations were associated with a significantly increased active pyruvate dehydrogenase complex in liver and skeletal muscle in each of the conditions examined. In liver, dichloroacetate fully activated (greater than 85%) the pyruvate dehydrogenase complex under all conditions. In skeletal muscle from chronic septic animals, the dichloroacetate-induced increases in active pyruvate dehydrogenase were significantly less than those observed in non-septic animals. The data suggest that although dichloroacetate can partially reverse the sepsis-induced effects on skeletal muscle pyruvate dehydrogenase activity, there may be additional regulatory factors in skeletal muscle from septic animals. The dichloroacetate stimulation of the pyruvate dehydrogenase activity may provide a pharmacological method for reducing the elevated lactate concentrations observed in chronic severe sepsis.

Abscess↗
PubMed1986

Pyruvate decarboxylating action of L-cycloserine. The significance of this in understanding its metabolic inhibitory action.

We present evidence which demonstrates that L-cycloserine, structural analog of L-alanine, which is known to be an effective aminotransferase inhibitor, is also a potent inhibitor of cellular pyruvate metabolism. This effect was found to be related to its almost instantaneous action in decreasing pyruvate concentrations in a dose-dependent manner. 1H nuclear magnetic resonance studies clearly demonstrate that the irreversible removal of pyruvate induced by L-cycloserine is caused by the decarboxylating action of the latter. Pyruvate disappearance induced by L-cycloserine can be stoichiometrically accounted for as acetate. The process does not involve any chemically detected transformation of L-cycloserine. These observations lead to two main considerations regarding the known action of L-cycloserine. First, its inhibitory effect on gluconeogenesis from lactate could be explained only on the basis of its ability to reduce pyruvate availability with no apparent need for transaminase inhibition. Second, its ability as a transaminase inhibitor should be reconsidered in view of its potent decarboxylating action on pyruvate and probably other oxoacids.

Animals↗
PubMed1986

A comparison between effects of pyruvate and herb medicines in preventing experimental oxalate urolithiasis in rats.

Sodium pyruvate, choreito (a herbal preparation), and urajirogashi (a herb) were added to a calcium-oxalate lithogenic diet (a glycolic-acid diet) to determine their effects in preventing lithogenicity. Male Wistar-strain rats which had been fed the glycolic-acid diet developed marked urinary calculi within 4 weeks. Rats in the groups fed a pyruvate diet had, however, almost no stones in the urinary system. The choreito and urajirogashi were slightly less effective than the pyruvate. Urinary oxalate excretion was high in all the groups during the experiment, especially in the pyruvate and the glycolic-acid groups, but, it was relatively lowered in the herb groups, especially towards the end of the experiment (p less than 0.05). Urinary citrate excretion was high in the pyruvate group, but it was significantly low in the other groups. In the choreito group, remarkable increases in urinary volume and magnesium excretion were observed; however, they were statistically non-significant and urinary calcium excretion was higher than in the glycolic-acid group during the experiment. Therefore, it can be concluded that choreito and urajirogashi may have some beneficial effect though any such effect is inferior to that of pyruvate, in preventing calculi formation, partly by decreasing the urinary oxalate excretion; increased urine volume and magnesium excretion may also have some other, additional effects in the choreito group.

Animals↗
PubMed1978

Determination of pyruvate oxidation rate and citric acid cycle activity in intact human leukocytes and fibroblasts.

We measured pyruvate oxidation in intact leukocytes and fibroblasts by measuring 14C02 production. The optimal pyruvate concentration appeared to be higher than that usually applied. Activities remained constant during the incubation and were proportional to the amount of tissue protein added. Man values (+/-SD) were 2.8 +/- 0.9 nmol/h per 10(6) cells and 37 +/- 14 nmol/h per mg of protein for leukocytes and fibroblasts, respectively, for [1-14C]pyruvate oxidation; and 2.1 +/- 0.8 nmol/h per 10(6) cells and 18 +/- 7 nmol/h per mg of protein, respectively for [2-14C]pyruvate oxidation. We compared oxidation rates of pyruvate and 2-oxoglutarate by intact cells with those of isolated mitochondria. The ratio of 14CO2 production vs. activity of mitochondrial marker enzyme demonstrated that the rate of pyruvate oxidation can adequately be assayed in intact cells, but that the permeability of the cell membrane is rate-limiting in the oxidation of 2-oxoglutarate. No significant oxidation of other intermediates of the citric acid cycle was found, presumably owing to a low rate of transport of these substances across the cell membrane.

Biological Transport↗
PubMed1983

Effects of insulin on CO2 fixation in adipose tissue. Evidence for regulation of pyruvate transport.

Insulin was found to double the rate of incorporation of H14CO3- into protein by segments of rat epididymal adipose tissue provided the incubation medium contained a suitable energy substrate such as fructose. Overall protein synthesis was increased by insulin to a lesser extent, one-third as measured by tritiated water indicating that insulin also increased CO2 fixation into amino acids. The latter could be demonstrated only when the tissue amino acid pools were expanded by the addition of aspartate to the incubation medium. The pattern of labeling observed in the amino acids indicated that CO2 fixation occurred primarily at the pyruvate carboxylase step. Addition of pyruvate to the incubation medium also increased CO2 fixation and this effect was not additive with that of insulin, suggesting that insulin acted by increasing the availability of pyruvate to the carboxylase. No change in carboxylase activity could be measured. Mitochondria isolated from tissue exposed to insulin retained a higher capacity to fix CO2 into acid-soluble products provided they were not freeze-thawed or sonicated. Uptake of pyruvate by mitochondria incubated 1 min at 2 degrees C or 5 s at 15 degrees C was doubled by prior insulin treatment of the tissue. It is concluded that insulin increases the flux through pyruvate carboxylase in adipose tissue in part by increasing the transport of pyruvate through the inner mitochondrial membrane.

Adipose Tissue↗
PubMed1983

Transport of pyruvate in mitochondria from different tumor cells.

A comparative study of the transport of pyruvate in mitochondria isolated from normal rat liver and from three tumors has been carried out. The Km for net pyruvate uptake in mitochondria isolated from Ehrlich ascites tumor cells is practically equal to that measured in normal rat liver mitochondria while, on the other hand, it is higher in Morris hepatomas 44 and 3924A. The Vmax of pyruvate uptake is depressed in all three types of tumor mitochondria as compared to that in the rat liver mitochondria, with the depression being higher in Morris hepatoma 3924A mitochondria. The lower activity of pyruvate translocator in mitochondria isolated from tumor cells as compared to that in rat liver mitochondria is also shown by depression of the rate of pyruvate-supported oxygen uptake. The results document a decreased activity of the pyruvate translocator in tumor mitochondria which seems to be correlated with the growth rate of the tumor cells.

Animals↗
PubMed1995

An amino acid substitution in the pyruvate dehydrogenase E1 alpha gene, affecting mitochondrial import of the precursor protein.

A mutation in the mitochondrial targeting sequence was characterized in a male patient with X chromosome-linked pyruvate dehydrogenase E1 alpha deficiency. The mutation was a base substitution of G by C at nucleotide 134 in the mitochondrial targeting sequence of the PDHA1 gene, resulting in an arginine-to-proline substitution at codon 10 (R10P). Pyruvate dehydrogenase activity in cultured skin fibroblasts was 28% of the control value, and immunoblot analysis revealed a decreased level of pyruvate dehydrogenase E1 alpha immunoreactivity. Chimeric constructs in which the normal and mutant pyruvate dehydrogenase E1 alpha targeting sequences were attached to the mitochondrial matrix protein ornithine transcarbamylase were synthesized in a cell free translation system, and mitochondrial import of normal and mutant proteins was compared in vitro. The results show that ornithine transcarbamylase targeted by the mutant pyruvate dehydrogenase E1 alpha sequence was translocated into the mitochondrial matrix at a reduced rate, suggesting that defective import is responsible for the reduced pyruvate dehydrogenase level in mitochondria. The mutation was also present in an affected brother and the mildly affected mother. The clinical presentations of this X chromosome-linked disorder in affected family members are discussed. To our knowledge, this is the first report of an amino acid substitution in a mitochondrial targeting sequence resulting in a human genetic disease.

Amino Acids↗