Membrane‐Bound c ‐Type Cytochromes in Heliobacillus mobilis

Abstract

The spectral and electrochemical parameters, as well as the orientations of the heme planes with respect to the membrane plane, of the c-type hemes present in membrane fragments from Heliobacillus mobilis were characterised by optical and EPR spectroscopy. Cytochrome cSs3 was thereby shown to represent at least four and possibly five heme species with the following characteristics: Em = -60 mV 2 10 mV, g, = 2.92, 60"; Em = +90 mV ? 10 mV, g , = 2.92, 90"; Em = +120 mV 2 20 mV, g, = 3.03 ; and Em = +170 mV 2 20 mV, g, = 3.03. The latter component may correspond to two hemes with redox midpoint potentials of Em = + 160 mV 2 20 mV and Em = +180 mV 2 20 mV (all E,,, values at pH 7.0). For the heme species having g, peaks at p 3 . 0 3 , determination of individual orientations was precluded due to the superposition of several differently oriented hemes. About one copy of each heme was found to be present per photosynthetic reaction centre, with the exception of the + 120 mV component for which a stoichioinetry of 2 hemesheaction centre was obtained. The heme proteins were detergent-solubilised and partially purified. Three c-type cytochromes that migrated with apparent molecular masses of 18, 29 and 50 kDa were detected on SDSPAGE. Optical redox titrations at pH 7.0 showed redox midpoint potentials of +160 mV? 10 mV for the 18-kDa cytochrome, and -60 mV? 10 mV, with possible contributions around +I60 mV, for the 50-kDa cytochrome. A tentative attribution of heme species observed in membranes to the isolated heme proteins is presented. The results obtained on H. mobilis are compared with those reported for green sulphur bacteria.

Heliobacterial photosynthesis

“…The gene sequences of the two other proteins in this complex are unknown. If the detergent used in this purification is non-ionic, the cytochrome c 553 complex is associated with yellowish-green pigments (Nitschke et al 1996). If the cytochrome c 553 complex is purified with an ionic detergent, the color is reddish-brown, consistent with the presence of a cytochrome (Albert et al 1998).…”

“…The 29 kDa heme binding protein is associated with the cytochrome bc complex (Nitschke et al 1995), which, in addition to cytochrome b, contains a Rieske Fe/S cluster Kramer et al 1997). The 50 and 18 kDa heme binding proteins have been proposed to represent the c-type cytochromes involved with forward donation to P798 + (Nitschke et al 1996). The gene for the 18 kDa cytochrome c protein has been sequenced, and has been named petJ (Albert et al 1998).…”

“…All the three cytochromes may donate electrons to P798 + either directly or indirectly (Nitschke et al 1995(Nitschke et al , 1996Kramer et al 1997). In whole cells, a cytochrome c donates to the primary donor in 100-600 ls following a laser flash (Vos et al 1989).…”

Heliobacterial Rieske/cytb complex

“…The globular, heme-containing domain of cytochrome c 553 is in the periplasmic space. A linker sequence of about 40 residues, with a peculiar amino acid composition (12 Ala, 7 Ser, 6 Lys, 6 Pro, 5 Glu, 2 Asn, 1 Gln) between the cysteine residue attached to the lipids and the globular domain, confers mobility to the protein (Schoepp et al 2000) evidenced by the absence of a preferential orientation of the g z signal of the heme of cytochrome c 553 in partially ordered membrane multilayers (Nitschke et al 1996).…”

“…The menaquinone pool was titrated to -60 mV (Kramer et al 1997), and the cytochrome c 553 to about ?180 mV (Nitschke et al 1996). Cytochrome c 553 reduces the primary donor of the RCI which has a midpoint potential of ?240 mV (Nitschke et al 1995).…”