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The electron transport system of Alcaligenes eutrophus H16

II. Respiratory activities and effect of specific inhibitors

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Abstract

In a previous work (Kömen et al. 1991) it has been concluded that membrane fragments isolated from autotrophically grown Alcaligenes eutrophus H16 contain several iron-sulphur centres along with haems of a-, b-, c-, and d-type. These redox components have been proposed to be part of a branched respiratory chain leading to multiple membrane bound oxidases. Here, some of the respiratory activities catalyzed by membrane fragments from wild type cells of A. eutrophus (H16) and, for comparison, Paracoccus denitrificans, have been investigated through the use of electron transport inhibitors. Cyanide (CN-) titration curves indicated that in A. eutrophus H16 oxidation of succinate and H2 preferentially proceeds via the cytochrome c oxidase(s) branch (I 50=2 · 10-5 M) whereas the NADH dependent respiration started being inhibited at higher CN- concentrations (I 50=5 · 10-4 M). In membranes isolated from both, cells harvested at late growth-phase (OD 12) and from a mutant deficient in cytochrome c oxidase activity (A. eutrophus RK1), respiration was insensitive to low CN- concentrations (< 10-4 M), and it was sustained by the high catalytic activities of two quinol oxidases. These alternative oxidases of b- (formally o-) and d-type showed different sensitivities to KCN (I 50=10-3 M and 10-2 M, respectively). Interestingly, the cytochrome c oxidase(s) dependent respiration of H16 membranes was insensitive to antimycin A but largely inhibited by myxothiazol (10-6 M). This, and previous work (Kömen et al. 1991), suggest that although the respiratory chain of A. eutrophus is endowed with a putative bc 1 complex, its biochemical nature and role in respiration of this organism are apparently different from those of P. denitrificans. The peculiarity of the respiratory chain of A. eutrophus is confirmed by the rotenone insensitivity of the NADH oxidation in both protoplasts and membrane fragments from wild type and soluble hydrogenase deficient cells (HF14 and HF160). A tentative model of the respiratory chain of autotrophically grown A. eutrophus is presented.

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Abbreviations

CTAB:

Cetyltrimethylammonium bromide

cytochrome c-551:

number: wavelength in nm; cytochrome c 270, number: mid-point potential in mV

DCIP:

2,6-dichlorophenol-indophenol

DMSO:

dimethyl sulfoxide

E m, 7 :

mid-point potential of an oxidation-reduction couple at pH 7.0

I 50 :

concentration of the inhibitor at which 50% inhibition of the activity is observed

KP-buffer:

potassium phosphate-buffer

OD:

optical density at 436 nm, 1 cm light path

PMS:

phenazine methosulfate

Q:

ubiquinone

Qi :

Q-reducing side of the bc 1 complex

Qo :

Q-oxidizing side of the bc 1 complex

TMPD:

N,N,N′,N′-tetramethyl-p-phenylenediamine

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Authors and Affiliations

  1. Institut für Mikrobiologie der Georg-August, Universität Göttingen, Grisebachstrasse 8, W-3400, Göttingen, Germany

    Ralf Kömen & Karin Schmidt

  2. Department of Biology, University of Bologna, I-40126, Bologna, Italy

    Davide Zannoni

Authors
  1. Ralf Kömen
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  2. Davide Zannoni
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  3. Karin Schmidt
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Kömen, R., Zannoni, D. & Schmidt, K. The electron transport system of Alcaligenes eutrophus H16. Arch. Microbiol. 155, 436–443 (1991). https://doi.org/10.1007/BF00244958

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  • DOI: https://doi.org/10.1007/BF00244958

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