TAILIEUCHUNG - Báo cáo Y học: Functional integration of mitochondrial and hydrogenosomal ADP/ATP carriers in the Escherichia coli membrane reveals different biochemical characteristics for plants, mammals and anaerobic chytrids

The expression of mitochondrial and hydrogenosomal ADP/ATP carriers (AACs) from plants, rat and the anaerobic chytridiomycete fungus Neocallimastix spec. L2 in Escherichia coli allows a functional integration of the recombinant proteins into the bacterial cytoplasmic membrane. For AAC1 and AAC2 from rat, apparent Km values of about 40 lM for ADP, and 105 lM or 140 lM, respectively, for ATP have been determined, similar to the data reported for isolated rat mitochondria. The apparent Km for ATP decreased up to 10-fold in the presence of the protonophore m-chlorocarbonylcyanide phenylhydrazone (CCCP). The hydrogenosomal AAC isolated from the chytrid fungus Neocallimastix spec | Eur. J. Biochem. 269 3172-3181 2002 FEBS 2002 doi Functional integration of mitochondrial and hydrogenosomal ADP ATP carriers in the Escherichia coh membrane reveals different biochemical characteristics for plants mammals and anaerobic chytrids Ilka Haferkamp1 Johannes H. P. Hackstein2 Frank G. J. Voncken2 Guillaume Schmit1 and Joachim Tjaden1 1 Pflanzenphysiologie Universitat Kaiserslautern Germany department of Evolutionary Microbiology Faculty of Science Catholic University of Nijmegen the Netherlands The expression of mitochondrial and hydrogenosomal ADP ATP carriers AACs from plants rat and the anaerobic chytridiomycete fungus Neocallimastix spec. L2 in Escherichia coli allows a functional integration of the recombinant proteins into the bacterial cytoplasmic membrane. For AAO add ACC2 from ten. apparent Km values of about 40 M for ADP and 105 M or 140 M respectively for ATP have been determined similar to the data reported for isolated rat mitochondria. The appoeant Km for ATP decreased up to 10-fold in the presence of the protonophore m-chlorocarbonyl-cyanide phenylhydrpzone CCCP . Tire l yrFghanosomlll AAC isolated from the chytrid fungus Neocallimastix spec. 22 exhibited the some chaaatOerisaics but the affinities for ADP 165 M and ATP mM were significantly lower. NorUalily. ACC1-3 from Arabidopsis thaliana and AAC1 from Solanum tuberosum potato showed significantly higher external affinities for both nucleotides 10-22 M they were only slightly influenced by CCCP. Studies on intact plant mitochondria confirmed these observations. Back exfaane 6x 10111 vhh 11111X1 1 E. coli cells expressing AACs indicate a preferential export of ATP for all AACs tested. This ÍS the lint extort of a functional integration of proteins belonging to the mitochondrial carrier family MCF into a bacterial cytoplasmic membrane. The tachnique diaoribtd hore provtdes a reta--ively simple and highly reproducible method for functional studies of .

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