Publications

• Purification, characterisation and reconstitution of glutaconyl-CoA decarboxylase, a biotin-dependent sodium pump from anaerobic bacteria.

  Buckel W., Semmler R.

  Glutaconyl-CoA decarboxylase from Acidaminococcus fermentans is a biotin enzyme, which is integrated into membranes. It is activated by Triton X-100 and inhibited by avidin. The results obtained by a combination of both agents indicate that biotin and the substrate-binding site are located on the ... >> More

  Glutaconyl-CoA decarboxylase from Acidaminococcus fermentans is a biotin enzyme, which is integrated into membranes. It is activated by Triton X-100 and inhibited by avidin. The results obtained by a combination of both agents indicate that biotin and the substrate-binding site are located on the same side of the membrane. The decarboxylase was solubilized with Triton X-100 and purified by affinity chromatography on monomeric avidin-Sepharose. The enzyme is composed of three types of polypeptides: the group of alpha chains (Mr 120000-140000) containing the biotin, the beta chain (60000) and an apparently hydrophobic gamma chain (35000). Sodium ions specifically protected the latter chain from tryptic digestion. It was supposed, therefore, that this chain might function as the Na+ channel. The beta and gamma chains but not the alpha chain could be labelled by N-ethyl-[14C]maleimide. Similar decarboxylases but with much smaller biotin peptides (Mr 15000-20000) were isolated from Peptococcus aerogenes and Clostridium symbiosum. The decarboxylases from all three organisms could be reconstituted to active sodium pumps by incubation with phospholipid vesicles and octylglucoside followed by dilution. The Na+ uptake catalysed by the enzyme from A. fermentans was completely inhibited by monensin and activated twofold by valinomycin/K+ indicating an electrogenic Na+ pump. The coupling between Na+ transport and decarboxylation was not tight. During the reaction the ratio decreased from initially 1 to 0.2. The three organisms mentioned above and Clostridium tetanomorphum without glutaconyl-CoA decarboxylase are able to ferment glutamate and require 10 mM Na+ for rapid growth. There is no correlation between the concentration of monensin necessary to inhibit growth and the presence of decarboxylase in these organisms. << Less

  Eur J Biochem 136:427-434(1983) [PubMed] [EuropePMC]

• Sodium ion-translocating decarboxylases.

  Buckel W.

  The review is concerned with three Na(+)-dependent biotin-containing decarboxylases, which catalyse the substitution of CO(2) by H(+) with retention of configuration (DeltaG degrees '=-30 kJ/mol): oxaloacetate decarboxylase from enterobacteria, methylmalonyl-CoA decarboxylase from Veillonella parv ... >> More

  The review is concerned with three Na(+)-dependent biotin-containing decarboxylases, which catalyse the substitution of CO(2) by H(+) with retention of configuration (DeltaG degrees '=-30 kJ/mol): oxaloacetate decarboxylase from enterobacteria, methylmalonyl-CoA decarboxylase from Veillonella parvula and Propiogenium modestum, and glutaconyl-CoA decarboxylase from Acidaminococcus fermentans. The enzymes represent complexes of four functional domains or subunits, a carboxytransferase, a mobile alanine- and proline-rich biotin carrier, a 9-11 membrane-spanning helix-containing Na(+)-dependent carboxybiotin decarboxylase and a membrane anchor. In the first catalytic step the carboxyl group of the substrate is converted to a kinetically activated carboxylate in N-carboxybiotin. After swing-over to the decarboxylase, an electrochemical Na(+) gradient is generated; the free energy of the decarboxylation is used to translocate 1-2 Na(+) from the inside to the outside, whereas the proton comes from the outside. At high [Na(+)], however, the decarboxylases appear to catalyse a mere Na(+)/Na(+) exchange. This finding has implications for the life of P. modestum in sea water, which relies on the synthesis of ATP via Delta(mu)Na(+) generated by decarboxylation. In many sequenced genomes from Bacteria and Archaea homologues of the carboxybiotin decarboxylase from A. fermentans with up to 80% sequence identity have been detected. << Less

  Biochim Biophys Acta 1505:15-27(2001) [PubMed] [EuropePMC]