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Energization of Bacillus subtilis membrane vesicles increases catalytic activity of succinate: Menaquinone oxidoreductase

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Abstract

In this work, high ΔμH+-dependent succinate oxidase activity has been demonstrated for the first time with membrane vesicles isolated from Bacillus subtilis. The maximal specific rate of succinate oxidation by coupled inside-out membrane vesicles isolated from a B. subtilis strain overproducing succinate:menaquinone oxidoreductase approaches the specific rate observed with the intact cells. Deenergization of the membrane vesicles with ionophores or alamethicin brings about an almost complete inhibition of succinate oxidation. An apparent K m for succinate during the energy-dependent succinate oxidase activity of the vesicles (2.2 mM) is higher by an order of magnitude than the K m value measured for the energy-independent reduction of 2,6-dichlorophenol indophenol. The data reveal critical importance of ΔμH+ for maintaining active electron transfer by succinate:menaquinone oxidoreductase. The role of ΔμH+ might consist in providing energy for thermodynamically unfavorable menaquinone reduction by succinate by virtue of transmembrane electron transport within the enzyme down the electric field; alternatively, ΔμH+ could play a regulatory role by maintaining the electroneutrally operating enzyme in a catalytically active conformation.

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Abbreviations

CCCP:

carbonyl cyanide m-chlorophenylhydrazone

DCCD:

N,N′-dicyclohexylcarbodiimide

DCPIP:

dichlorophenol indophenol

FAD:

flavin adenine dinucleotide

PMS:

phenazine methosulfate

QFR:

quinol:fumarate oxidoreductase

SQR:

succinate:quinone oxidoreductase

TMPD:

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

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Authors and Affiliations

  1. Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992, Moscow, Russia

    N. V. Azarkina & A. A. Konstantinov

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  1. N. V. Azarkina
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  2. A. A. Konstantinov
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Correspondence to A. A. Konstantinov.

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Published in Russian in Biokhimiya, 2010, Vol. 75, No. 1, pp. 63–77.

Originally published in Biochemistry (Moscow) On-Line Papers in Press, as Manuscript BM09-162, December 20, 2009.

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Azarkina, N.V., Konstantinov, A.A. Energization of Bacillus subtilis membrane vesicles increases catalytic activity of succinate: Menaquinone oxidoreductase. Biochemistry Moscow 75, 50–62 (2010). https://doi.org/10.1134/S0006297910010074

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  • DOI: https://doi.org/10.1134/S0006297910010074

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