Protein Self-Assembly and Lipid Binding in the Folding of the Potassium Channel KcsA

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• 作者：Francisco N. Barrera ; M. Lourdes Renart ; Jos&eacute ; A. Poveda ; Ben de Kruijff ; J. Antoinette Killian ; Jos&eacute ; M. Gonzá ; lez-Ros
• 刊名：Biochemistry
• 出版年：2008
• 出版时间：February 19, 2008
• 年：2008
• 卷：47
• 期：7
• 页码：2123 - 2133
• 全文大小：204K
• 年卷期：v.47,no.7(February 19, 2008)
• ISSN：1520-4995

文摘

Moderate concentrations of the alcohol 2,2,2-trifluoroethanol (TFE) cause the coupled unfoldingand dissociation into subunits of the homotetrameric potassium channel KcsA, in a process that is partiallyirreversible when the protein is solubilized in plain dodecyl -D-maltoside (DDM) micelles [Barrera et al.(2005) Biochemistry 44, 14344-52]. Here we report that the transition from the folded tetramer to theunfolded monomer becomes completely reversible when KcsA is solubilized in mixed micelles composedof the detergent DDM and the lipids DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine) and DOPG(1,2-dioleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)]). This result suggests that lipids may act as effectorsin the tetramerization of KcsA. The observed reversibility allowed the determination of the standard freeenergy of the folding reaction of KcsA: G = 30.5 ± 3.1 kcal·mol^-1. We also observed that, prior to theunfolding of the tetramer, the presence of lower TFE concentrations causes the disassembly ofsupramolecular clusters of KcsA into the individual tetrameric molecules. Within the limits of experimentalresolution, this is also a reversible process, but unlike the tetramer to monomer transition from above, thelevel of clustering is not influenced by the presence of solubilized lipids. These observations suggest adistinct role of the lipids in the different in vitro assembly steps (folding/tetramerization and clustering)of KcsA.