Vasilev, C. orcid.org/0000-0002-0536-882X, Mayneord, G.E., Brindley, A.A. et al. (2 more authors) (2019) Dissecting the cytochrome c(2)–reaction centre interaction in bacterial photosynthesis using single molecule force spectroscopy. Biochemical Journal, 476 (15). pp. 2173-2190. ISSN 0264-6021
Abstract
The reversible docking of small, diffusible redox proteins onto a membrane protein complex is a common feature of bacterial, mitochondrial and photosynthetic electron transfer (ET) chains. Spectroscopic studies of ensembles of such redox partners have been used to determine ET rates and dissociation constants. Here, we report a single-molecule analysis of the forces that stabilise transient ET complexes. We examined the interaction of two components of bacterial photosynthesis, cytochrome c2 and the reaction centre (RC) complex, using dynamic force spectroscopy and PeakForce quantitative nanomechanical imaging. RC–LH1–PufX complexes, attached to silicon nitride AFM probes and maintained in a photo-oxidised state, were lowered onto a silicon oxide substrate bearing dispersed, immobilised and reduced cytochrome c2 molecules. Microscale patterns of cytochrome c2 and the cyan fluorescent protein were used to validate the specificity of recognition between tip-attached RCs and surface-tethered cytochrome c2. Following the transient association of photo-oxidised RC and reduced cytochrome c2 molecules, retraction of the RC-functionalised probe met with resistance, and forces between 112 and 887 pN were required to disrupt the post-ET RC–c2 complex, depending on the retraction velocities used. If tip-attached RCs were reduced instead, the probability of interaction with reduced cytochrome c2 molecules decreased 5-fold. Thus, the redox states of the cytochrome c2 haem cofactor and RC ‘special pair’ bacteriochlorophyll dimer are important for establishing a productive ET complex. The millisecond persistence of the post-ET cytochrome c2[oxidised]–RC[reduced] ‘product’ state is compatible with rates of cyclic photosynthetic ET, at physiologically relevant light intensities.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2019 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (https://creativecommons.org/licenses/by/4.0/). |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) > Department of Molecular Biology and Biotechnology (Sheffield) |
Funding Information: | Funder Grant number Biotechnology and Biological Sciences Research Council (BBSRC) BB/M000265/1; BB/P002005/1 |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 25 Sep 2019 14:01 |
Last Modified: | 26 Sep 2019 01:32 |
Status: | Published |
Publisher: | Portland Press Ltd. |
Refereed: | Yes |
Identification Number: | 10.1042/bcj20170519 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:151340 |