Abstract
Functional reintegration into lipid environments represents a major challenge for in vitro investigation of integral membrane proteins (IMPs). Here, we report a new approach, termed LMNG Auto-insertion Reintegration (LAiR), for reintegration of IMPs into lipid bilayers within minutes. The resulting proteoliposomes displayed an unprecedented capability to maintain proton gradients and long-term stability. LAiR allowed for monitoring catalysis of a membrane-bound, physiologically relevant polyisoprenoid quinone substrate by Escherichia coli cytochromes bo3 (cbo3) and bd (cbd) under control of the proton motive force. LAiR also facilitated bulk-phase detection and physiological assessment of the “proton leak” in cbo3, a controversial catalytic state that previously was only approachable at the single-molecule level. LAiR maintained the multisubunit integrity and higher-order oligomeric states of the delicate mammalian F-ATP synthase. Given that LAiR can be applied to both liposomes and planar membrane bilayers and is compatible with IMPs and lipids from prokaryotic and eukaryotic sources, we anticipate LAiR to be applied broadly across basic research, pharmaceutical applications, and biotechnology.
Original language | English |
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Pages (from-to) | 494-507 |
Number of pages | 14 |
Journal | ACS Central Science |
Volume | 9 |
Issue number | 3 |
Early online date | 22 Feb 2023 |
DOIs | |
Publication status | Published - 22 Mar 2023 |
Bibliographical note
Funding Information:This work was supported by a long-term International fellowship from the Japan Society for the Promotion of Sciences (JSPS; P14383) to D.G.G.M.; Delft University of Technology Start-up Grant to D.G.G.M.; a BINDS grant from AMED (JP16K07266 to Atsunori Oshima and C.G., JP22ama121001j0001 to Masaki Yamamoto and C.G.); a Grants-in-Aid for Scientific Research (B) (JP 17H03647) from MEXT to C.G.; the International Joint Research Promotion Program from Osaka University to Genji Kurisu and C.G.; a JST-CREST Grant Number (JP18071859) to K.M.; the Naito Foundation Subsidy for Female Researchers after Maternity Leave (C.J.) and JSPS 25–5370 (C.J.). A.H.A. wishes to thank the Royal Embassy of Saudi Arabia (NL) and King Abdulaziz University in Saudi Arabia for financial support.
Publisher Copyright:
© 2023 The Authors. Published by American Chemical Society.
Funding
This work was supported by a long-term International fellowship from the Japan Society for the Promotion of Sciences (JSPS; P14383) to D.G.G.M.; Delft University of Technology Start-up Grant to D.G.G.M.; a BINDS grant from AMED (JP16K07266 to Atsunori Oshima and C.G., JP22ama121001j0001 to Masaki Yamamoto and C.G.); a Grants-in-Aid for Scientific Research (B) (JP 17H03647) from MEXT to C.G.; the International Joint Research Promotion Program from Osaka University to Genji Kurisu and C.G.; a JST-CREST Grant Number (JP18071859) to K.M.; the Naito Foundation Subsidy for Female Researchers after Maternity Leave (C.J.) and JSPS 25–5370 (C.J.). A.H.A. wishes to thank the Royal Embassy of Saudi Arabia (NL) and King Abdulaziz University in Saudi Arabia for financial support.
Funders | Funder number |
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Osaka University to Genji Kurisu | |
King Abdulaziz University | |
Technische Universiteit Delft | |
Ministry of Education, Culture, Sports, Science and Technology | |
Japan Society for the Promotion of Science | P14383, 22KJ1638 |
Japan Agency for Medical Research and Development | JP 17H03647, JP16K07266, JP22ama121001j0001 |
Core Research for Evolutional Science and Technology | JP18071859 |
Naito Foundation | 25–5370 |