Heteronuclear multidimensional NMR and homology modelling studies of the C-terminal nucleotide-binding domain of the human mitochondrial ABC transporter ABCB6.
Human ATP-binding cassette, sub-family B, member 6 (ABCB6) is a mitochondrial ABC transporter, and presumably contributes to iron homeostasis. Aimed at understanding the structural basis for the conformational changes accompanying the substrate-transportation cycle, we have studied the C-terminal nucleotide-binding domain of ABCB6 (ABCB6-C) in both the nucleotide-free and ADP-bound states by heteronuclear multidimensional NMR and homology modelling. A non-linear sampling scheme was utilised for indirectly acquired 13C and 15N dimensions of all 3D triple-resonance NMR experiments, in order to overcome the instability and the low solubility of ABCB6-C. The backbone resonances for approximately 25% of non-proline residues, which are mostly distributed around the functionally important loops and in the Helical domain, were not observed for nucleotide-free form of ABCB6-C. From the pH, temperature and magnetic field strength dependencies of the resonance intensities, we concluded that this incompleteness in the assignments is mainly due to the exchange between multiple conformations at an intermediate rate on the NMR timescale. These localised conformational dynamics remained in ADP-bound ABCB6-C except for the loops responsible for adenine base and alpha/beta-phosphate binding. These results revealed that the localised dynamic cooperativity, which was recently proposed for a prokaryotic ABC MJ1267, also exists in a higher eukaryotic ABC, and is presumably shared by all members of the ABC family. Since the Helical domain is the putative interface to the transmembrane domain, this cooperativity may explain the coupled functions between domains in the substrate-transportation cycle.
Kurashima-Ito
Kaori
K
Cellular and Molecular Biology Laboratory, RIKEN, Wako, Saitama, 351-0198, Japan.
Ikeya
Teppei
T
Senbongi
Hiroshi
H
Tochio
Hidehito
H
Mikawa
Tsutomu
T
Shibata
Takehiko
T
Ito
Yutaka
Y
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D006801:Humans; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D040681:Structural Homology, Protein
C107991:ABCB6 protein, human; D018528:ATP-Binding Cassette Transporters; D024101:Mitochondrial Proteins; D000244:Adenosine Diphosphate