ATP hydrolysis-dependent conformational changes in the extracellular domain of ABCA1 are associated with apoA-I binding.
ATP-binding cassette protein A1 (ABCA1) plays a major role in cholesterol homeostasis and high-density lipoprotein (HDL) metabolism. Although it is predicted that apolipoprotein A-I (apoA-I) directly binds to ABCA1, the physiological importance of this interaction is still controversial and the conformation required for apoA-I binding is unclear. In this study, the role of the two nucleotide-binding domains (NBD) of ABCA1 in apoA-I binding was determined by inserting a TEV protease recognition sequence in the linker region of ABCA1. Analyses of ATP binding and occlusion to wild-type ABCA1 and various NBD mutants revealed that ATP binds equally to both NBDs and is hydrolyzed at both NBDs. The interaction with apoA-I and the apoA-I-dependent cholesterol efflux required not only ATP binding but also hydrolysis in both NBDs. NBD mutations and cellular ATP depletion decreased the accessibility of antibodies to a hemagglutinin (HA) epitope that was inserted at position 443 in the extracellular domain (ECD), suggesting that the conformation of ECDs is altered by ATP hydrolysis at both NBDs. These results suggest that ATP hydrolysis at both NBDs induces conformational changes in the ECDs, which are associated with apoA-I binding and cholesterol efflux.
Laboratory of Cellular Biochemistry, Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University , Kyoto 606-8502, Japan.
Kohjiro
K
Nagao
Kei
K
Takahashi
Yuya
Y
Azuma
Mie
M
Takada
Yasuhisa
Y
Kimura
Michinori
M
Matsuo
Noriyuki
N
Kioka
Kazumitsu
K
Ueda
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D057809:HEK293 Cells; D006388:Hemagglutinins / Q000276:immunology; D006801:Humans; D006868:Hydrolysis; D008239:Lysine / Q000235:genetics; D051379:Mice; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D051381:Rats
C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D006388:Hemagglutinins; D000255:Adenosine Triphosphate; D002784:Cholesterol; D008239:Lysine