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. 2019 Nov 27;9:17662. doi: 10.1038/s41598-019-54198-8

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Plasmid borne nhaA expresses a mixture of NhaA monomers and unstable dimers in BKT12, a mutant strain that does not synthesize cardiolipin. TA16/pAXH3 cells expressing WT-NhaA and a mutant that does not synthesize cardiolipin (BKT12/pI^Q, pAXH3) bearing pAXH3 and pI^Q (a plasmidic combination needed for NhaA IPTG control expression) were grown on minimal medium to 0.8 OD₆₀₀, IPTG-induced for 2 h. Then, high-pressure membranes were isolated, NiNTA affinity-purified proteins were isolated, and samples (6 µg protein/7.5 µL) were mixed with equal volumes of native gel sampling buffer, titrated to final pH 7, incubated in the indicated DDM concentrations for 10 min at 23 °C, and resolved on native gel. The experiment was conducted three times with identical results.