Electron transfer proteins from the haloalkaliphilic archaeon Natronobacterium pharaonis: possible components of the respiratory chain include cytochrome bc and a terminal oxidase cytochrome ba3.

Abstract

Natronobacterium pharaonis, an aerobic haloalkaliphilic archaebacterium, expresses high concentrations of redox proteins as do alkaliphilic eubacteria. The first redox protein characterized from N. pharaonis was halocyanin [Scharf, B., & Engelhard, M. (1993) Biochemistry 32, 12894-12900], a small blue copper protein. It is a peripheral membrane protein and is conjectured to function in a manner similar to plastocyanin. In the present work, the respiratory chain is further elucidated and the purification and characterization of the most abundant components cytochrome bc and cytochrome ba3 from the membrane fraction are described. The cytochrome bc complex consists of a 14 and an 18 kDa subunit in a 1:1 ratio, with heme c bound to the larger polypeptide. An Fe-S subunit similar to that found in eukaryotic bc complexes has not yet been identified. The second membrane complex carries two different heme groups of the ba3-type as well as copper. It contains two subunits of 36 and 40 kDa. This cytochrome ba3 binds carbon monoxide, a feature common to terminal oxidases. There is no spectroscopic evidence for a second terminal oxidase; hence, under the growth conditions chosen the respiratory chain of N. pharaonis appears to be unbranched. In addition to these cytochromes, a succinate dehydrogenase which is solubilized from the membrane by detergents was isolated. A cytochrome c which was isolated from the cytosol has an unusually high molecular weight and a redox potential of -142 mV. A second cytosolic protein, ferredoxin, was purified to homogeneity. A comparison of the redox potentials of the isolated proteins with those obtained from the native membrane allows the construction of a possible electron transfer chain.