Heme centers of Rhodothermus marinus respiratory chain. Characterization of its cbb3 oxidase.

Abstract

Rhodothermus (R.) marinus, a thermohalophilic gram-negative, and strict aerobic bacterium, has a rather distinct respiratory chain, containing a caa3 terminal oxidase, a novel cytochrome bc complex and a HiPIP, which is an electron carrier between this complex and a terminal oxidase (Pereira et al (1999a, c). To further elucidate this unusual respiratory system, its membrane-bound heme centers were characterized by visible and EPR spectroscopies as well as by redox potentiometry. Rhodothermus marinus contains mostly B- and C-type hemes; a small amount of A-type heme is also detected. The heme centers have relatively low reduction potentials, ranging from ca. +250 to -60 mV, at pH 7. A Rieske-type center was not detected, suggesting the absence of a canonical complex III. The major terminal oxidase expressed by R. marinus is a cbb3-type oxidase. Its presence is in agreement with molecular biology studies, which reveal the existence of a gene encoding for a FixN-type oxidase. The oxidase was partially purified and appears to have five subunits, with apparent molecular masses of 64, 57, 36, 26 (C-type heme subunit), and 13 kDa. It contains two low-spin heme C centers, one high-, and one low-spin heme B centers. A full description of the equilibrium redox behavior of the heme centers was obtained for a cbb3 oxidase for the first time. The optical spectrum for each heme center and the corresponding reduction potentials were determined at pH 7: + 195 (heme C), +120 (heme B), -50 (heme C), and -50 mV (heme B3).