How curved membranes recruit amphipathic helices and protein anchoring motifs

Abstract

Lipids and several specialized proteins are thought to be able to sense the curvature of membranes (MC). Here we used quantitative fluorescence microscopy to measure curvature-selective binding of amphipathic motifs on single liposomes 50–700 nm in diameter. Our results revealed that sensing is predominantly mediated by a higher density of binding sites on curved membranes instead of higher affinity. We proposed a model based on curvature-induced defects in lipid packing that related these findings to lipid sorting and accurately predicted the existence of a new ubiquitous class of curvature sensors: membrane-anchored proteins. The fact that unrelated structural motifs such as α-helices and alkyl chains sense MC led us to propose that MC sensing is a generic property of curved membranes rather than a property of the anchoring molecules. We therefore anticipate that MC will promote the redistribution of proteins that are anchored in membranes through other types of hydrophobic moieties.

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Figure 1: A fluorescence-based assay to measure MC-selective binding of biomolecules on single liposomes.
Figure 2: Quantification of the parameters governing the MC-selective binding of two amphipathic motifs, eAH and aAH, on single liposomes.
Figure 3: Model explaining MC sensing by amphiphilic molecules at saturating concentration.
Figure 4: MC sensing by alkyl chains.
Figure 5: MC sensing by proteins carrying alkyl chain membrane anchors.
Figure 6: Dependence of binding affinity and Bmax on MC for double palmitoylated GST.

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Acknowledgements

We thank H. Wennerstrom, T. Heimburg and T. Bjornholm for critically reading the manuscript; J.-B. Perez and K.L. Martinez for help with data treatment; N. Kirkby (University of Copenhagen Hospital, Rigshospitalet, Department of Clinical Microbiology) for providing palmitoylated ovalbumin; J.L. Baneres (Institut des Biomolécules Max Mousseron, University of Montpellier) for the generous contribution of Gβ1γ2 and H. McMahon (MRC Laboratory of Molecular Biology) for kindly providing the plasmid for rat endophilin A1. This work was supported by the Danish Councils for Scientific and Strategic Research and partly by the European Union FP6–2004–IST–4 program NEMOSLAB.

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V.K.B. developed the assay; N.S.H. designed most experiments and recorded and treated most data with help from J.L., V.K.B., P.-Y.B. and J.C.; A.H.K. developed the image treatment; K.L.M. purified and labeled the AH and GST constructs; P.H. helped formulate the model; D.S. designed and supervised the project and wrote the main text together with N.S.H. and V.K.B. The manuscript was discussed and corrected by all co-authors.

Corresponding author

Correspondence to Dimitrios Stamou.

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Hatzakis, N., Bhatia, V., Larsen, J. et al. How curved membranes recruit amphipathic helices and protein anchoring motifs. Nat Chem Biol 5, 835–841 (2009). https://doi.org/10.1038/nchembio.213

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