Purification and partial characterization of two cytochrome oxidases (caa3 and o) from the thermophilic bacterium PS3.
Two cytochrome oxidases, cytochrome aa3 (EC 1.9.3.1) and cytochrome o, have been purified from the membranes of a thermophilic bacterium, PS3. The enzymes were solubilized with Triton X-100 and purified to apparent homogeneity on anion-exchange columns. The properties of the three-subunit cytochrome oxidase complex caa3 obtained here are compared with the same enzyme isolated by Sone, N. and Yanagita, Y. (1982) (Biochim. Biophys. Acta 682, 216-226). On storage, the purified caa3 enzyme undergoes denaturation; a shoulder at 432 nm seen in (CO-reduced)-minus-reduced difference spectra may be due in part to denaturation products of the enzyme. The purified cytochrome o is more stable. At room temperature, the reduced-minus-oxidized difference spectrum shows absorbance maxima at 427 and 559 nm; at 77 K, its alpha-band is split into 554 and 557 nm components. At room temperature, the CO-reduced-minus-reduced spectrum shows troughs at 430 nm and 560 nm. Dissociating polyacrylamide gel electrophoresis suggests that the purified cytochrome o is composed of one type of subunit with an apparent molecular mass of 47 000-48 000. Metal analysis of the purified enzyme demonstrated the lack of copper. Both oxidases, purified in the presence of Triton X-100, exist in highly polydisperse forms.
D001419:Bacteria / Q000201:enzymology*; D003576:Electron Transport Complex IV / Q000302:isolation & purification*; D006358:Hot Temperature; D046911:Macromolecular Substances; D008970:Molecular Weight; D013057:Spectrum Analysis
D046911:Macromolecular Substances; D003576:Electron Transport Complex IV