Conformational Change Induced by ATP Binding in the Multidrug ATP-Binding Cassette Transporter BmrA
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- 作者：Cé ; dric Orelle ; Francesca Gubellini ; Anne Durand ; Sergio Marco ; Daniel Lé ; vy ; Philippe Gros ; Attilio Di Pietro ; Jean-Michel Jault
- 刊名：Biochemistry
- 出版年：2008
- 出版时间：February 26, 2008
- 年：2008
- 卷：47
- 期：8
- 页码：2404 - 2412
- 全文大小：387K
- 年卷期：v.47,no.8(February 26, 2008)
- ISSN：1520-4995
文摘
ATP-binding cassette (ABC) transporters are involved in the transport of a wide variety ofsubstrates, and ATP-driven dimerization of their nucleotide binding domains (NBDs) has been suggestedto be one of the most energetic steps of their catalytic cycle. Taking advantage of the propensity ofBmrA, a bacterial multidrug resistance ABC transporter, to form stable, highly ordered ring-shapedstructures [Chami et al. (2002) J. Mol. Biol. 315, 1075-1085], we show here that addition of ATP in thepresence of Mg2+ prevented ring formation or destroyed the previously formed rings. To pinpoint thecatalytic step responsible for such an effect, two classes of hydrolysis-deficient mutants were furtherstudied. In contrast to hydrolytically inactive glutamate mutants that behaved essentially as the wild-type,lysine Walker A mutants formed ring-shaped structures even in the presence of ATP-Mg. Although thelatter mutants still bound ATP-Mg, and even slowly hydrolyzed it for the K380R mutant, they were mostlikely unable to undergo a proper NBD dimerization upon ATP-Mg addition. The ATP-driven dimerizationstep, which was still permitted in glutamate mutants and led to a stable conformation suitable to monitorthe growth of 2D crystals, appeared therefore responsible for destabilization of the BmrA ring structures.Our results provide direct visual evidence that the ATP-induced NBD dimerization triggers a conformationalchange large enough in BmrA to destabilize the rings, which is consistent with the assumption that thisstep might constitute the "power stroke" for ABC transporters.