The orientation of the three haems of the ‘in situ’ ubiquinol oxidase, cytochrome bd, of Escherichia coli

Abstract

The Escherichia coli cytochrome bd complex incorporates three haems as prosthetic groups. In the ferric form these are a predominantly high-spin chlorin (haem d), a high-spin haem b (b595) and a low-spin haem b (b558). The orientations of these three haems have been determined by e.p.r. studies on oriented multilayer preparations of cytoplasmic membrane fragments. The low-spin haem b (b558) and the high-spin haem d are oriented with their haem planes perpendicular to the membrane plane. The high-spin haem b595 is oriented with its haem plane at approx. 55 degrees to the membrane plane. A minor low-spin component, attributable to a low-spin subpopulation of the haem d, is also oriented with its haem plane perpendicular to the membrane plane.

The cytochrome bd respiratory oxygen reductases

“…Both subunits are required for the assembly of heme b 595 and heme d , suggesting that these two hemes may reside at the subunit interface [151]. Heme b 595 appears to be oriented with its heme plane at ~55° to the plane of the membrane [223]. The millimolar extinction coefficients used commonly for the determination of the cytochrome bd concentration in E. coli and A. vinelandii are listed in Table 2.…”

Proton pumping by NADH:ubiquinone oxidoreductase. A redox driven conformational change mechanism?

“…A redox midpoint potential dependence of 360 mV/pH unit around neutral pH values reported for bovine complex I [29] has been considered as an indication that cluster N2 may be directly involved in the proton translocation mechanism [7]. However, this attractive option seems unlikely now: For Y. lipolytica complex I we have determined a pK ox of V6 and a pK red of V7 for the protonable group associated with iron^sulfur cluster N2.…”

“…N1a is bound to the 24 kDa subunit [27,28]. In bovine complex I, this cluster has the lowest redox midpoint potential (E m;7 = 3370 mV) and exhibits a pH dependence of 360 mV/ pH [29,30]. Analysis of the isolated 24 kDa subunit from bovine mitochondria and di¡erent bacteria by protein-¢lm voltammetry revealed that at low ionic strength the reduction potential changes only by V100 mV between pH 5 and 9 [31].…”

Femtosecond resolution of ligand-heme interactions in the high-affinity quinol oxidase bd : A di-heme active site?

“…Such communication could involve (i) excitonic interactions between the hemes and (ii) steric interactions. Concerning the first mechanism, it can be calculated by using the point dipole approximation for excitonic coupling (37) that for two hemes at a distance of 7 Å with heme planes at 55° (38); an 8% change in heme d oscillator strength would be sufficient to alter the interaction with heme b 595 by Ϸ 60 cm Ϫ1 (a shift of Ϸ1 nm at 440 nm). As to the second mechanism, bound CO could be sandwiched between the two hemes and thus sterically affect the heme b 595 iron-macrocycle structure.…”