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Effects induced by rotenone during aerobic growth ofParacoccus denitrificans in continuous culture

Changes in energy conservation and electron transport associated with NADH dehydrogenase

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Abstract

Paracoccus denitrificans was grown aerobically in chemostat culture in the presence of rotenone. After 6 to 10 generation times, cells showed an oxygen uptake which was completely rotenone-insensitive after removal of rotenone by washing with bovine serum albumin containing medium.

The →H+/O ratio of these cells for endogenous substrates decreased from about 7.50 to 3.95. The latter ratio was similar to the value obtained for starved cells oxidizing exogenous succinate, indicating that site I phosphorylation was absent in these rotenone-insensitive cells.

Membrane particles prepared from these cells showed an 80% decrease in activity of reduced nicotinamide adenine dinucleotide oxidase and reduced nicotinamide adenine dinucleotide-ferricyanide oxidoreductase, while also the kinetic behaviour of the reduced nicotinamide adenine dinucleotide dehydrogenase in the reduced nicotinamide adenine dinucleotide-ferricyanide oxidoreductase assay was changed. Moreover the reduced nicotinamide adenine dinucleotide oxidase activity was practically rotenone-insensitive.

Electron paramagnetic resonance spectroscopy on membrane particles from rotenone-insensitive cells at 15 K revealed that the resonance lines atg z ≈ 2.05 andg y ≈g x ≈ 1.92 arising from iron-sulfur center 2 were undetectable. The intensities of the other electron paramagnetic resonance signals originating from reduced nicotinamide adenine dinucleotide dehydrogenase linked iron-sulfur centers were only slightly diminished.

These observations confirm our previous suggestion that site I phosphorylation, rotenone sensitivity and the presence of iron-sulfur center 2 are correlated.

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Abbreviations

EPR:

electron paramagnetic resonance

BSA:

bovine serum albumin

CCCP:

carbonylcyanide m-chlorophenylhydrazone

NAD:

nicotinamide adenine dinucleotide

NADP:

nicotinamide adenine dinucleotide phosphate

ATP:

adenosine triphosphate

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Authors and Affiliations

  1. Department of Microbiology, Biological Laboratory, Free University, De Boelelaan 1087, NL-1007 MC, Amsterdam, The Netherlands

    E. M. Meijer, M. G. Schuitenmaker, F. C. Boogerd & A. H. Stouthamer

  2. Laboratory of Biochemistry, B.C.P. Jansen Institute, University of Amsterdam, Plantage Muidergracht 12, Amsterdam, The Netherlands

    R. Wever

Authors
  1. E. M. Meijer
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  2. M. G. Schuitenmaker
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  3. F. C. Boogerd
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  4. R. Wever
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  5. A. H. Stouthamer
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Meijer, E.M., Schuitenmaker, M.G., Boogerd, F.C. et al. Effects induced by rotenone during aerobic growth ofParacoccus denitrificans in continuous culture. Arch. Microbiol. 119, 119–127 (1978). https://doi.org/10.1007/BF00964262

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