Abstract
The membranes of the thermoacidophilic archaeon Sulfolobus metallicus exhibit an oxygen consumption activity of 0.5 nmol O-2 min(-1) mg(-1), which is insensitive to rotenone, suggesting the presence of a type-II NADH dehydrogenase. Following this observation, the enzyme was purified from solubilised membranes and characterised. The pure protein is a monomer with an apparent molecular mass of 49 ...
Abstract
The membranes of the thermoacidophilic archaeon Sulfolobus metallicus exhibit an oxygen consumption activity of 0.5 nmol O-2 min(-1) mg(-1), which is insensitive to rotenone, suggesting the presence of a type-II NADH dehydrogenase. Following this observation, the enzyme was purified from solubilised membranes and characterised. The pure protein is a monomer with an apparent molecular mass of 49 kDa, having a high N-terminal amino acid sequence similarity towards other prokaryotic enzymes of the same type. It contains a covalently attached flavin, which was identified as being FMN by P-31-NMR spectroscopy, a novelty among type-II NADH dehydrogenases. Metal analysis showed the absence of iron, indicating that no Fes clusters are present in the protein. The average reduction potential of the FMN group was determined to be + 160 mV, at 25degreesC and pH 6.5, by redox titrations monitored by visible spectroscopy. Catalytically, the enzyme is a NADH:quinone oxidoreductase, as it is capable of transferring electrons from NADH to several quiriones, including ubiquinone-1, ubiquinone-2 and caldariella quinone. Maximal turnover rates of 195 mumol NADH oxidized min(-1) mg(-1) at 60degreesC were obtained using ubiquinone-2 as electron acceptor, after enzyme dilution and incubation with phospholipids. (C) 2002 Elsevier Science B.V. All rights reserved.
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