Caught in the Act: ATP hydrolysis of an ABC-multidrug transporter followed by real-time magic angle spinning NMR
Caught in the Act: ATP hydrolysis of an ABC-multidrug transporter followed by real-time magic angle spinning NMR
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DOI：
10.1016/j.febslet.2008.09.033
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发表时间：
2008-10-15
期刊：
影响因子：
3.5
通讯作者：
Glaubitz, Clemens
中科院分区：
文献类型：
--
作者：
Hellmich, Ute A.;Haase, Winfried;Glaubitz, Clemens
The ATP binding cassette (ABC) transporter LmrA from Lactococcus lactis transports cytotoxic molecules at the expense of ATP. Molecular and kinetic details of LmrA can be assessed by solid-state nuclear magnetic resonance (ssNMR), if functional reconstitution at a high protein-lipid ratio can be achieved and the kinetic rate constants are small enough. In order to follow ATP hydrolysis directly by P-31-magic angle spinning (MAS) nuclear magnetic resonance (NMR), we generated such conditions by reconstituting LmrA-dK388, a mutant with slower ATP turnover rate, at a protein-lipid ration of 1: 150. By analysing time-resolved P-31 spectra, protein activity has been directly assessed. These data demonstrate the general possibility to perform ssNMR studies on a fully active full length ABC transporter and also form the foundation for further kinetic studies on LmrA by NMR. (C) 2008 Federation of European Biochemical Societies. Published by Elsevier B. V. All rights reserved.
