Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.4.1.2 (acetyl-CoA carboxylase)
 2,876 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Acetyl-CoA carboxylase, a major rate-limiting enzyme for fatty acid synthesis, is subject to acute regulation by both allosteric modulation and covalent enzyme phosphorylation. Because citrate activation of the enzyme in vitro requires citrate concentrations far in excess of intracellular levels, we have attempted to identify other ligands which might mediate carboxylase activity. Heated liver extracts contain a potent endogenous activator of carboxylase assayed in dialyzed high speed liver supernatant; the activator elutes behind the salt volume of a Bio-Gel P-6 gel filtration column, is destroyed by alkaline phosphatase, and is adsorbed by charcoal. This activator activity is shared by several guanine nucleotides (5'-GTP, 5'-GDP, 5'-GMP, and 3':5'-cyclic GMP). Further separation of the endogenous activator by high pressure liquid chromatography reveals a carboxylase-activating compound which co-elutes with 5'-GMP. The guanine nucleotides are potent activators of carboxylase activity at intracellular nucleotide concentrations and permit expression of maximal enzyme velocity at cytosolic citrate concentrations. However, we have been unable to document any effects of guanine nucleotides on isolated rat liver acetyl-CoA carboxylase. While the mechanisms of these effects remain to be elucidated, they suggest that the guanine nucleotides may be important intracellular regulators of carboxylase activity and of fatty acid synthesis.
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PMID:Regulation of acetyl-CoA carboxylase by guanine nucleotides. 611 55

Definitive evidence is presented for the bifunctional nature of the biotin repressor protein which possesses both regulatory and enzymatic activities. The repressor protein can activate biotin in the presence of ATP to form biotinyl-5'-adenylate, the co-repressor which remains tightly bound to the repressor protein. This complex can either bind to the operator site and inhibit transcription or transfer the biotinyl moiety to a lysine residue of the apoenzyme of acetyl-CoA carboxylase. The two activities were coincident throughout a purification procedure which resulted in a 3500-fold increase in activity. Gel electrophoresis of the purified preparation, under native or denaturing conditions, showed three proteins with the activity corresponding to the major protein band of apparent Mr = 34,000. On gel exclusion chromatography, the activity was also associated with a protein of Mr varying fro 37,000-44,000, indicating the protein is monomeric. The occasional appearance of multiple bands with biological activity in the native gels suggests that the repressor protein can also exist in multimeric forms. On chromatofocusing, the repressor activity and the holoenzyme synthetase activity were coincidental, with the peak of activity at pH 7.2, the isoelectric point. Only a single protein band with Mr = 34,000 was observed on SDS gel electrophoresis of all fractions showing activity.
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PMID:Purification and properties of the biotin repressor. A bifunctional protein. 612 46

Protein phosphatase-2B was purified from extracts of rabbit skeletal muscle by a procedure that involved fractionation with ammonium sulphate, chromatography on DEAE-Sepharose, fractionation with poly(ethylene glycol), gel filtration on Sephadex G-200 (Mr = 98000 +/- 4000), chromatography on Affi-Gel Blue and affinity chromatography on calmodulin-Sepharose. The enzyme was purified 3500-fold in seven days with an overall yield of 0.5%. The alpha-subunit of phosphorylase kinase, protein phosphatase inhibitor-1 and the myosin P-light chain from rabbit skeletal muscle were dephosphorylated by protein phosphatase-2B with similar kinetic constants. The alpha-subunit of phosphorylase kinase was dephosphorylated at least 100-fold more rapidly than the beta-subunit, while glycogen phosphorylase, glycogen synthase, histones H1 and H2B, ATP-citrate lyase, acetyl-CoA carboxylase, L-pyruvate kinase and protein synthesis initiation factor eIF-2 were not dephosphorylated at significant rates. Protein phosphatase-2B became activated 10-fold by calmodulin (A0.5 = 6 nM) after chromatography on DEAE-Sepharose and this degree of activation was maintained throughout the remainder of the purification. Calmodulin increased the Vmax of the reaction without altering the Km for inhibitor-1. The activity of protein phosphatase-2B was completely dependent on Ca2+ in the presence or absence of calmodulin. Half-maximal activation was observed at 1.0 microM Ca2+ in the absence, and at 0.5 microM Ca2+ in the presence, of 0.03 microM calmodulin. Protein phosphatase-2B was inhibited completely by trifluoperazine; half-maximal inhibition occurred at 45 microM in the absence and 35 microM in the presence of 0.03 microM calmodulin. The metabolic role of protein phosphatase-2B in vivo is discussed in the light of the observation that this enzyme is probably identical to a major calmodulin-binding protein of neural tissue termed calcineurin or CaM-BP80 [Stewart, A. A., Ingebritsen, T. S., Manalan, A., Klee, C. B., and Cohen, P. (1982) FEBS Lett. 137, 80-84].
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PMID:The protein phosphatases involved in cellular regulation. 5. Purification and properties of a Ca2+/calmodulin-dependent protein phosphatase (2B) from rabbit skeletal muscle. 630 28

When two copies of the sequences spanning -57 to -35 of the fatty acid synthase (FAS) or -64 to -41 of the ATP citrate-lyase (ACL) gene linked to a reporter gene were transfected into primary cultured hepatocytes, the reporter activities significantly increased in response to insulin/glucose treatment. In cotransfection experiments of the FAS(-57/-35) with the Sp1 or Sp3 expression vector, the reporter activities of transcription were suppressed by Sp1 and stimulated by Sp3. In the cotransfection experiments of ACL(-64/-41), the activities were suppressed by Sp1 but were unchanged by Sp3. A similar effect of Sp1 and Sp3 on transcription was seen in mRNA concentrations and enzyme activities of endogenous FAS and ACL. Moreover, the mRNA concentrations and enzyme activities of endogenous acetyl-CoA carboxylase were suppressed by Sp1 and greatly increased by Sp3. Gel mobility super shift assays using antibodies against Sp1 or Sp3 revealed the binding of the transcription factors Sp1 and Sp3 with the GC rich regions located within FAS(-57/-35) and ACL(-64/-41) genes. The formation of DNA-protein complexes was decreased in rats fed a high-carbohydrate diet in comparison with that in fasted rats, but feeding the corn oil diet inhibited this decrease. In Western immunoblotting assay, however, the amount of Sp1 and Sp3 remained unchanged in the dietary conditions. Therefore, the binding of DNA-protein complexes was not due to changes in the amount of Sp1 and Sp3 but to changes in the binding activity, suggesting that these transcription factors may be an important determinant of lipogenic enzyme expression.
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PMID:Transcriptional regulation of fatty acid synthase gene and ATP citrate-lyase gene by Sp1 and Sp3 in rat hepatocytes(1). 1061 88

Acetyl-CoA carboxylase regulates the rate of fatty acid synthesis. This enzyme in plants is localized in plastids and is believed to be composed of biotin carboxyl carrier protein, biotin carboxylase, and carboxyltransferase made up of alpha and beta polypeptides, although the enzyme has not been purified yet. Accumulated evidence shows that pea plastidic acetyl-CoA carboxylase is activated by light and the activation is caused by light-dependent reduction of carboxyltransferase, but not of biotin carboxylase, via a redox cascade. To understand the reductive activation of carboxyltransferase at the molecular level here, we obtained the active enzyme composed of decahistidine-tagged (His tag) alpha and beta polypeptides through the expression of the pea plastidic carboxyltransferase gene in Escherichia coli. Gel filtration showed that the molecular size of the recombinant carboxyltransferase is in agreement with that of partially purified carboxyltransferase from pea chloroplasts. The catalytic activity of the recombinant enzyme was similar to that of native carboxyltransferase. These results indicate that the molecular structure and conformation of recombinant carboxyltransferase resemble those of its native counterpart and that native carboxyltransferase is indeed composed of alpha and beta polypeptides. This recombinant enzyme was activated by dithiothreitol, a known reductant of S-S bonds, with a profile similar to that of its native counterpart. The recombinant enzyme was activated by reduced thioredoxin-f, a signal transducer of redox potential in chloroplasts under irradiation. Thus, this enzyme was redox-regulated, like that of the native carboxyltransferase.
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PMID:Recombinant carboxyltransferase responsive to redox of pea plastidic acetyl-CoA carboxylase. 1074 68