TetL Tetracycline Efflux Protein from Bacillus subtilis Is a Dimer in the Membrane and in Detergent Solution

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• 作者：Markus Safferling ; Heather Griffith ; Jie Jin ; Josh Sharp ; Magdia De Jesus ; Caroline Ng ; Terry A. Krulwich ; and Da-Neng Wang
• 刊名：Biochemistry
• 出版年：2003
• 出版时间：December 2, 2003
• 年：2003
• 卷：42
• 期：47
• 页码：13969 - 13976
• 全文大小：136K
• 年卷期：v.42,no.47(December 2, 2003)
• ISSN：1520-4995

文摘

The TetL antiporter from the Bacillus subtilis inner membrane is a tetracycline-divalent cationefflux protein that is energized by the electrochemical proton gradient across the membrane. In this study,we expressed tetL in Escherichia coli and investigated the oligomeric state of TetL in the membrane andin detergent solution. Evidence for an oligomeric state of TetL emerged from SDS-PAGE and Westernblot analysis of membrane samples as well as purified protein samples from cells that expressed twodifferently tagged TetL species. Furthermore, no formation or restoration of TetL oligomers occurredupon detergent solubilization of the membrane. Rather, oligomeric forms established in vivo persistedafter solubilization. Mass spectrometry of the purified protein showed the absence of proteolysis andposttranslational modifications. Analytical size-exclusion chromatography of the purified protein revealeda dimeric TetL in dodecyl-maltoside solution. In addition, TetL dimers were found in a number of otherdetergents and over a wide pH range. It is therefore likely that the oligomeric form of the protein in themembrane is also a dimer.