Characterization of human ATP-binding cassette protein subfamily D reconstituted into proteoliposomes

• Published in: Biochemical and biophysical research communications 2018-02, Vol.496 (4), p.1122-1127
• Main Authors: Okamoto, Takumi, Kawaguchi, Kosuke, Watanabe, Shiro, Agustina, Rina, Ikejima, Toshiki, Ikeda, Keisuke, Nakano, Minoru, Morita, Masashi, Imanaka, Tsuneo
• Format: Article
• Language: English
• Subjects: ABC protein subfamily D; Acyl-CoA thioesterase; ATPase; Proteoliposome; Purification
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2018.01.153

In mammals, four ATP-binding cassette (ABC) proteins belonging to subfamily D have been identified. ABCD1‒3 are located on peroxisomal membrane and play an important role in the transportation of various fatty acid-CoA derivatives, including very long chain fatty acid-CoA, into peroxisomes. ABCD4 is located on lysosomal membrane and is suggested to be involved in the transport of vitamin B12 from lysosomes to the cytosol. However, the precise transport mechanism by which these ABC transporters facilitate the import or export of substrate has yet to be well elucidated. In this study, the overexpression of human ABCD1‒4 in the methylotrophic yeast Pichia pastoris and a purification procedure were developed. The detergent-solubilized proteins were reconstituted into liposomes. ABCD1‒4 displayed stable ATPase activity, which was inhibited by AlF3. Furthermore, ABCD1‒4 were found to possess an equal levels of acyl-CoA thioesterase activity. Proteoliposomes is expected to be an aid in the further biochemical characterization of ABCD transporters. •Human ABC protein subfamily D, hABCD1, 2, 3 and 4 were successfully expressed in Pichia pastoris.•hABCD1‒4 were purified and reconstituted into liposomes as an active form.•hABCD1‒4 were shown to possess nearly equal acyl-CoA thioesterase activity.