SULFOLOBUS-ACIDOCALDARIUS TERMINAL OXIDASE - KINETIC INVESTIGATION AND ITS STRUCTURAL INTERPRETATION

The thermoacidophilic archaebacterium Sulfolobus acidocaldarius posses ses a very unusual terminal oxidase. We report original kinetic experi ments on membranes of this microorganism carried out by stopped flow u sing time-resolved optical spectroscopy combined with singular value d ecomposition analysis. The reduced-oxidized kinetic difference spectru m of the Sulfolobus membranes is characterized by three significant pe aks in the visible region at 605, 586, and 560 nm. The 605-nm peak and part of the 586-nm peak (cytochrome aa(3)-type quinol oxidase) are re duced synchronously by both ascorbate plus N,N,N',N'-tetramethyl-p-phe nylendiamine (TMPD) and dithionite, and they are very rapidly oxidized by molecular oxygen. A second pool of cytochromes seems to contribute to the 586-nm peak which is not reduced by ascorbate plus TMPD and re acts very slowly with dithionite. The b-type cytochromes (560 nm peak) are reduced by both reductants and are essentially ''non-autoxidizabl e'' at room temperature. Only one CO binding site with spectral featur es, kinetic properties, and ligand affinity not very dissimilar from t hose of mammalian cytochrome oxidase can be detected in the ascorbate- reduced membranes. On the contrary, a second CO binding site having un usual properties for aa(3) terminal oxidases can be detected in the di thionite-reduced membranes.