A large Ca2+-dependent channel formed by recombinant ADP/ATP carrier from Neurospora crassa resembles the mitochondrial permeability transition pore

Nikolai Broustovetski, Maximilian Tropschug, Simone Heimpel, Doerthe Heidkämper, Martin Klingenberg

Research output: Contribution to journalArticle

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Abstract

Strong support for the central role of the ADP/ATP carrier (AAC) in the mitochondrial permeability transition (mPT) is provided by the single-channel current measurements in patch-clamp experiments with the isolated reconstituted AAC. In previous work [Brustovetsky, N., and Klingenberg, M. (1996) Biochemistry 35, 8483-8488], this technique was applied to the AAC isolated from bovine heart mitochondria. Here we used recombinant AAC (rAAC) from Neurospora crassa expressed in E. coli, since AAC from mammalian sources cannot be expresssed in E. coli. The rAAC is free from residual mitochondrial components which might associate with the AAC in preparation from bovine heart. Ca2+-dependent channels with up to 600 pS are obtained, which are gated at > 150 mV. The channel corresponds to a preferential matrix-outside orientation of rAAC in the patch membrane as shown with carboxyatractylate and a polar gating asymmetry. The channel is inhibited by ADP and bongkrekate, not by carboxyatractylate. Cyclophilin, isolated from Neurospora crassa, suppresses the gating, thus increasing conductivity at high positive voltage. Cyclosporin A abolishes the cyclophilin effect. ADP does not eliminate the cyclophilin effect but produces fast large-amplitude flickering of the channel without a stable decrease of the channel conductance. Also the pro-oxidant tert-butyl hydroperoxide reversibly suppresses voltage gating of the channel. The results show that the AAC can be a conducting component of the mPT pore, exhibiting similar characteristics as the mPT pore (response to Ca2+, BKA, ADP), with a cyclophilin and pro-oxidant-sensitive gating at high voltage.

Original languageEnglish (US)
Pages (from-to)11804-11811
Number of pages8
JournalBiochemistry
Volume41
Issue number39
DOIs
StatePublished - Oct 1 2002
Externally publishedYes

Fingerprint

ATP Translocases Mitochondrial ADP
Neurospora crassa
Cyclophilins
Adenosine Diphosphate
Escherichia coli
Reactive Oxygen Species
Bongkrekic Acid
Electric potential
Flickering
tert-Butylhydroperoxide
Heart Mitochondria
Biochemistry
Mitochondria
Clamping devices
Electric current measurement
mitochondrial permeability transition pore
Cyclosporine
Permeability
Membranes

ASJC Scopus subject areas

  • Biochemistry

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A large Ca2+-dependent channel formed by recombinant ADP/ATP carrier from Neurospora crassa resembles the mitochondrial permeability transition pore. / Broustovetski, Nikolai; Tropschug, Maximilian; Heimpel, Simone; Heidkämper, Doerthe; Klingenberg, Martin.

In: Biochemistry, Vol. 41, No. 39, 01.10.2002, p. 11804-11811.

Research output: Contribution to journalArticle

Broustovetski, Nikolai ; Tropschug, Maximilian ; Heimpel, Simone ; Heidkämper, Doerthe ; Klingenberg, Martin. / A large Ca2+-dependent channel formed by recombinant ADP/ATP carrier from Neurospora crassa resembles the mitochondrial permeability transition pore. In: Biochemistry. 2002 ; Vol. 41, No. 39. pp. 11804-11811.
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