Purification and characterization of the reconstitutively active adenine nucleotide carrier from mitochondria of Jerusalem artichoke (Helianthus tuberosus L.) tubers.

摘要: The adenine nucleotide carrier from Jerusalem artichoke (Helianthus Tuberosus L.) tubers mitochondria was solubilized with Triton X-100 and purified by sequential chromatography on hydroxapatite Matrex Gel Blue B in the presence of cardiolipin asolectin. SDS gel electrophoresis fraction showed a single polypeptide band an apparent molecular mass 33 kDa. When reconstituted liposomes, catalyzed pyridoxal 5′-phosphate-sensitive ATP/ATP exchange. It 75-fold recovery 15% protein yield 0.18% respect to mitochondrial extract. Among various substrates inhibitors tested, transported only ATP, ADP, GTP inhibited bongkrekate, phenylisothiocyanate, 5′-phosphate, mersalyl p-hydroxymercuribenzoate (but not N-ethylmaleimide). Atractyloside carboxyatractyloside (at concentrations normally inhibitory animal plant mitochondria) were without effect mitochondria. V max exchange determined be 0.53 μmol/min per mg at 25°C. half-saturation constant K m corresponding inhibition K i 20.4 μM for ATP 45 ADP. activation energy 28 KJ/mol between 5 30°C. N-terminal amino acid partial sequence homology ANT maize shoots.