Biochemical characterisation of the isolated Anc2 adenine nucleotide carrier from Saccharomyces cerevisiae mitochondria.
The yeast mitochondrial adenine nucleotide carrier isoform encoded by the ANC2 gene has been specifically expressed in a yeast strain disrupted for the two other genes, ANC1 and ANC3. Isolation of the carrier in a functional form was achieved by utilisation of a mixture of two detergents, dodecylmaltoside and Emulphogen. The intrinsic fluorescence of the Anc2 protein was specifically and rapidly enhanced upon addition of the transportable nucleotides ADP and ATP. Fluorescence enhancement was prevented or reversed by the addition of a stoichiometric amount of CATR. Addition of CATR alone elicited a dose-dependent decrease of fluorescence. The ANC2-specific yeast stain offers the means to study a single ADP/ATP carrier, with a well-defined amino acid sequence, suitable for analysis of substrate- or inhibitor-induced conformational changes.
Brandolin
G
G
UA 1130 du CNRS, Département de Biologie Moléculaire et Structurale, Grenoble, France.
Le Saux
A
A
Trezeguet
V
V
Vignais
P V
PV
Lauquin
G J
GJ
D001278:Atractyloside / Q000031:analogs & derivatives / Q000378:metabolism; D001665:Binding Sites; D005453:Fluorescence; D008928:Mitochondria / Q000201:enzymology*; D000226:Mitochondrial ADP, ATP Translocases / Q000302:isolation & purification / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D001278:Atractyloside; D000226:Mitochondrial ADP, ATP Translocases; C003853:carboxyatractyloside