Structure of AMP-PNP-bound vitamin B12 transporter BtuCD-F
Structure of AMP-PNP-bound vitamin B12 transporter BtuCD-F
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DOI：
10.1038/nature11442
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发表时间：
2012-10-18
期刊：
影响因子：
64.8
通讯作者：
Locher, Kaspar P.
中科院分区：
文献类型：
--
作者：
Korkhov, Vladimir M.;Mireku, Samantha A.;Locher, Kaspar P.
ATP结合盒（ABC）转运蛋白BtuCD介导维生素B-12穿过大肠杆菌内膜的摄取。以前的结构显示了载脂蛋白状态的构象，但运输机制仍然不清楚。在这里，我们报告的3.5埃晶体结构的转运蛋白结合蛋白复合物BtuCD-BtuF（BtuCD-F）被困在β-γ-亚氨基腺苷5 '-磷酸（AMP-PNP）结合的中间状态。虽然ABC结构域（BtuD亚基）形成预期的封闭夹心二聚体，跨膜BtuC亚基采用新的构象，与中央易位途径密封由以前未识别的细胞质门。因此，完全封闭的腔形成在膜的大约一半处。它足够大以容纳维生素B-12分子，并且放射性配体捕获显示脂质体重构的BtuCD-F在AMP-PNP存在下确实含有结合的B-12。结合工程二硫化物交联和功能测定，我们的数据表明了一种意想不到的蠕动转运机制，与其他ABC转运蛋白中观察到的机制不同。
The ATP-binding cassette (ABC) transporter BtuCD mediates the uptake of vitamin B-12 across the inner membrane of Escherichia coli. Previous structures have shown the conformations of apo states, but the transport mechanism has remained unclear. Here we report the 3.5 angstrom crystal structure of the transporter-binding protein complex BtuCD-BtuF (BtuCD-F) trapped in an beta-gamma-imidoadenosine 5'-phosphate (AMP-PNP)-bound intermediate state. Although the ABC domains (BtuD subunits) form the expected closed sandwich dimer, the membrane-spanning BtuC subunits adopt a new conformation, with the central translocation pathway sealed by a previously unrecognized cytoplasmic gate. A fully enclosed cavity is thus formed approximately halfway across the membrane. It is large enough to accommodate a vitamin B-12 molecule, and radioligand trapping showed that liposome-reconstituted BtuCD-F indeed contains bound B-12 in the presence of AMP-PNP. In combination with engineered disulphide crosslinking and functional assays, our data suggest an unexpected peristaltic transport mechanism that is distinct from those observed in other ABC transporters.