Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.1.2.13 (aldolase)
 3,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

E. coli pyruvate oxidase (pyruvate:ferricytochrome b1 oxidoreductase, EC 1.2.2.2) is a peripheral membrane flavoenzyme which has been purified to homogeneity. In vivo the oxidase resides on the inner surface of the cytoplasmic membrane and is coupled to the bacterial electron transport chain. In vitro, the purified oxidase requires lipids for full enzymatic activity. Previous studies have characterized the conformational and energetic coupling between the lipid-binding site(s) and the catalytic active site. The affinity of the enzyme for phospholipids and detergents is significantly enhanced when the flavoprotein is in the reduced form, i.e., in the presence of pyruvate and the required cofactor, thiamin pyrophosphate. The lipid-binding studies were hindered due to the complicating factor of the self-association of the substrate-reduced flavoprotein. In this paper, fluorescence techniques are employed to measure the binding of a detergent-like activator to the oxidase. The experiments are performed at much lower protein concentrations than previously employed, so that protein aggregation is not a problem. The chromophore on the activator, 2-(N-decyl)aminonaphthalene-6-sulfonic acid is effective at quenching the pyruvate oxidase intrinsic tryptophan fluorescence. Quenching titrations are used to obtain the binding isotherm. AT DNS concentrations less than 10(-5) M, the results show a larger amount of DNS binding to the reduced flavoprotein than to the oxidized form of the enzyme. This is the concentration range where DNS is an effective activator of the enzyme. This represents a class of binding sites specifically found on pyruvate oxidase and not apparent in other proteins such as lysozyme or aldolase. At the DNS concentration which is optimum for activation approx. 20 molecules of DNS are bound per enzyme tetramer in the absence of the substrate. The pyruvate-reduced form of the enzyme binds about 40--50 molecules of DNS per tetramer. Qualitatively, the results are similar to what was previously found for both sodium dodecyl sulfate and cetyl trimethylammonium bromide. However, in both these cases, the amount of bound detergent was nearly an order of magnitude less than the values obtained using DNS.
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PMID:The binding of a fluorescent activator 2-(N-decyl)aminonaphthalene-6-sulfonic acid to pyruvate oxidase. 700 Jan 89

Proteomic techniques were used to identify cardiac proteins from whole heart homogenate and heart mitochondria of Fisher 344/Brown Norway F1 rats, which suffer protein nitration as a consequence of biological aging. Soluble proteins from young (5 mo old) and old (26 mo old) animals were separated by one- and two-dimensional gel electrophoresis. One- and two-dimensional Western blots with an anti-nitrotyrosine antibody show an age-related increase in the immunoresponse of a few specific proteins, which were identified by nanoelectrospray ionization-tandem mass spectrometry (NSI-MS/MS). Complementary proteins were immunoprecipitated with an immobilized anti-nitrotyrosine antibody followed by NSI-MS/MS analysis. A total of 48 proteins were putatively identified. Among the identified proteins were alpha-enolase, alpha-aldolase, desmin, aconitate hydratase, methylmalonate semialdehyde dehydrogenase, 3-ketoacyl-CoA thiolase, acetyl-CoA acetyltransferase, GAPDH, malate dehydrogenase, creatine kinase, electron-transfer flavoprotein, manganese-superoxide dismutase, F1-ATPase, and the voltage-dependent anion channel. Some contaminating blood proteins including transferrin and fibrinogen beta-chain precursor showed increased levels of nitration as well. MS/MS analysis located nitration at Y105 of the electron-transfer flavoprotein. Among the identified proteins, there are important enzymes responsible for energy production and metabolism as well as proteins involved in the structural integrity of the cells. Our results are consistent with age-dependent increased oxidative stress and with free radical-dependent damage of proteins. Possibly the oxidative modifications of the identified proteins contribute to the age-dependent degeneration and functional decline of heart proteins.
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PMID:Proteomic identification of 3-nitrotyrosine-containing rat cardiac proteins: effects of biological aging. 1534 82

Although 4-hydroxy-2-nonenal (HNE, a product of lipid peroxidation) is a major cause of oxidative damage inside skeletal muscles, the exact proteins modified by HNE are unknown. We used two-dimensional electrophoresis, immunoblotting, and mass spectrometry to identify selective proteins targeted by HNE inside the diaphragm of rats under two conditions: severe sepsis [induced by E. coli lipopolysaccharides (LPS)] and during strenuous muscle contractions elicited by severe inspiratory resistive loading (IRL). Diaphragm HNE-protein adduct formation (detected with a polyclonal antibody) increased significantly after 1 and 3 h of LPS injection with a return to baseline values thereafter. Similarly, HNE-protein adduct formation inside the diaphragm rose significantly after 6 but not 3 h of IRL. Mass spectrometry analysis of HNE-modified proteins revealed enolase 3b, aldolase and triosephosphate isomerase 1, creatine kinase, carbonic anyhdrase III, aconitase 2, dihydrolipoamide dehydrogenase, and electron transfer flavoprotein-beta. Measurements of in vitro enolase activity in the presence of pure HNE revealed that HNE significantly attenuated enolase activity in a dose-dependent fashion, suggesting that HNE-derived modifications have inhibitory effects on enzyme activity. We conclude that lipid peroxidation products may inhibit muscle contractile performance through selective targeting of enzymes involved in glycolysis, energy production as well as CO(2) hydration.
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PMID:Modifications of proteins by 4-hydroxy-2-nonenal in the ventilatory muscles of rats. 1660 97

The content and distribution of body lipids are of special interest for production efficiency and meat quality in the farm animal industry. Triglycerides represent the most variable fraction of tissue lipids, and are mainly stored in adipocytes. Although several studies have reported regional differences in the expression of genes and their products in adipocytes from various species, the characteristics of i.m. adipocytes remain poorly described. To evaluate adipocyte features according to muscle and other fat locations, adipocyte proteins were isolated from trapezius skeletal muscle, and intermuscular, s.c., or perirenal adipose tissues from 6 female pigs (80 d of age). Protein extracts were labeled and analyzed by 2-dimensional, fluorescent, differential gel electrophoresis. The comparisons revealed that 149 spots were always differentially expressed (P < 0.05, ratio exceeding |2|-fold difference) between i.m. adipocytes and the fat cells derived from the 3 other adipose locations. The proteins that were downregulated in i.m. fat cells belonged to various metabolic pathways, such as lipogenesis (cytosolic malate dehydrogenase and isocitrate dehydrogenase, P < 0.01), glycolysis (enolases and aldolase, P </= 0.01), lipolysis (perilipin, P < 0.01), fatty acid oxidation (long-chain fatty-acyl CoA dehydrogenase, P < 0.01), and energy transfer (catalase, voltage-dependent anion channel 1, and electron-transfer flavoprotein, P < 0.05). In contrast, both prohibitin-1 and cell division cycle 42 homolog, with possible roles in cell growth, were up-regulated (P < 0.05) in i.m. adipocytes compared with other fat cells. Fewer differences were observed when adipocytes isolated from s.c., perirenal, and intermuscular fat tissues were compared, with a maximum of 17 spots differing significantly in abundance between perirenal and s.c. adipose tissues. The findings that proteins involved in both anabolic and energy-yielding catabolic pathways are downregulated in i.m. adipocytes compared with s.c., visceral, or intermuscular adipocytes, suggest that the metabolic activity of i.m. adipocytes is low. Thus, triggering adipogenesis rather than cell metabolism per se might be a valuable strategy to control lipid deposition in pig skeletal muscles.
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PMID:Regional differences in porcine adipocytes isolated from skeletal muscle and adipose tissues as identified by a proteomic approach. 1831 Apr 87

X-linked muscular dystrophy is a highly progressive disease of childhood and characterized by primary genetic abnormalities in the dystrophin gene. Senescent mdx specimens were used for a large-scale survey of potential age-related alterations in the dystrophic phenotype, because the established mdx animal model of dystrophinopathy exhibits progressive deterioration of muscle tissue with age. Since the mdx tibialis anterior muscle is a frequently used model system in muscular dystrophy research, we employed this particular muscle to determine global changes in the dystrophic skeletal muscle proteome. The comparison of mdx mice aged 8 weeks versus 22 months by mass-spectrometry-based proteomics revealed altered expression levels in 8 distinct protein species. Increased levels were shown for carbonic anhydrase, aldolase, and electron transferring flavoprotein, while the expressions of pyruvate kinase, myosin, tropomyosin, and the small heat shock protein Hsp27 were found to be reduced in aged muscle. Immunoblotting confirmed age-dependent changes in the density of key muscle proteins in mdx muscle. Thus, segmental necrosis in mdx tibialis anterior muscle appears to trigger age-related protein perturbations due to dystrophin deficiency. The identification of novel indicators of progressive muscular dystrophy might be useful for the establishment of a muscle subtype-specific biomarker signature of dystrophinopathy.
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PMID:Profiling of age-related changes in the tibialis anterior muscle proteome of the mdx mouse model of dystrophinopathy. 2309 55