OCCURRENCE OF 2 PLASTIDIC ATP ADP TRANSPORTERS IN ARABIDOPSIS-THALIANA L - MOLECULAR CHARACTERIZATION AND COMPARATIVE STRUCTURAL-ANALYSIS OF SIMILAR ATP/ADP TRANSLOCATORS FROM PLASTIDS AND RICKETTSIA-PROWAZEKII/

Recently, we sequenced a cDNA clone from Arabidopsis thaliana L. encod ing an ATP/ADP transporter protein (AATP1) located in the plastid enve lope membrane. The deduced amino acid sequence of AATP1 exhibits a hig h degree of similarity (>66%) to the ATP/ADP transporter from the obli gate intracellular gram-negative bacterium Rickettsia prowazekii. Here we report a second plastidic ATP/ADP carrier from A. thaliana (AATP2) . As deduced from the amino acid sequence, AATP2 exhibits 77.6% identi ty to AATP1 and 36% to the rickettsial protein. Hydropathy analysis in dicates that all three translocators are highly hydrophobic membrane p roteins, which exhibit marked similarities and differences. The AATP1 translocator lacks the sixth transmembrane domain that is present in A ATP2 and the bacterial transporter in R. prowazekii. In contrast to AA TP1 and the bacterial transport protein, only AATP2 exhibits a truncat ed C-terminal end. To compare the general biochemical properties of AA TP2 with the known transport properties of AATP1 we cloned the entire AATP2 cDNA into plasmid pJT118, leading to the presence of an addition al N-terminal histidine tag of 10 amino acids. For heterologous expres sion of His(10)-AATP2 we chose the Escherichia coli strain C43, which was reported recently to allow overproduction of eucaryotic membrane t ransport proteins. After transformation and subsequent induction by is opropylthio-2-D-galactopyranoside intact E. coli cells harbouring plas mid pJT118 showed import of radioactively labelled ATP and ADP. As ded uced from a Lineweaver-Burk analysis His(10)-AATP2 exhibited apparent K-m values for ATP and ADP of 22 mu M and 20 mu M, respectively. Impor t of ADP into His(10)-AATP2-expressing E. coli cells occurred at a rat e of 24 nmol.mg protein(-1).h(-1), which was about threefold faster th an import of ATP These biochemical characteristics are similar to tran sport properties of the heterologously expressed His(10)-AATP1 protein .