Characterization of the ATP transporter in the reconstituted rough endoplasmic reticulum proteoliposomes.
Adenosine triphosphate (ATP) transporter from rat liver rough endoplasmic reticulum (RER) was solubilized and reconstituted into phosphatidylcholine liposomes. The RER proteoliposomes, resulting from optimizing some reconstitution parameters, had an apparent K(m) value of 1.5 microM and a V(max) of 286 pmol min(-1) (mg protein)(-1) and showed higher affinity for ATP and a lower V(max) value than intact RER (K(m) of 6.5 microM and V(max) of 1 nmol). ATP transport was time- and temperature-dependent, inhibited by 4, 4'-diisothiocyanostilbene-2,2'-disulfonic acid, which is known as an inhibitor of anion transporters including ATP transporter, but was not affected by atractyloside, a specific inhibitor of mitochondrial ADP/ATP carrier. The internal and external effects of various nucleotides on the ATP transport were examined. ATP transport was cis-inhibited strongly by ADP and weakly by AMP. ADP-preloaded RER proteoliposomes showed a specific increase of ATP transport activity while AMP-preloaded RER proteoliposomes did not show the enhanced overshoot peak in the ATP uptake plot. These results demonstrate the ADP/ATP antiport mechanism of ATP transport in rat liver RER.
Shin
S J
SJ
Center for Molecular Catalysis, Department of Chemistry, Seoul National University, Shillim-dong, San 56-1, Kwanak-ku, 151-742, Seoul, South Korea.
Lee
W K
WK
Lim
H W
HW
Park
J
J
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D011510:Proteolipids / Q000737:chemistry*; D051381:Rats; D017208:Rats, Wistar
D018528:ATP-Binding Cassette Transporters; D011510:Proteolipids; C020485:proteoliposomes; D000255:Adenosine Triphosphate