Publication: Sticholysin, Sphingomyelin, and Cholesterol: A Closer Look at a Tripartite Interaction
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Publication Date
2019-06-18
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Cell Press
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Abstract
Actinoporins are a group of soluble toxic proteins that bind to membranes containing sphingomyelin (SM) and oligomerize to form pores. Sticholysin II (StnII) is a member of the actinoporin family, produced by Stichodactyla helianthus. Cholesterol (Chol) is known to enhance the activity of StnII. However, the molecular mechanisms behind this activation have remained obscure, although the activation is not Chol specific but rather sterol specific. To further explore how bilayer lipids affect or are affected by StnII, we have used a multiprobe approach (fluorescent analogs of both Chol and SM) in combination with a series of StnII tryptophan (Trp)-mutants, to study StnII/bilayer interactions. First we compared StnII bilayer permeabilization in the presence of Chol or oleoyl-ceramide (OCer). The comparison was done since both Chol and OCer have a 1-hydroxyl which help to orient the molecule in the bilayer (although OCer have additional polar functional groups). Both Chol and OCer also have increased affinity for SM, which StnII may recognize. However, our results show that only Chol was able to activate StnII-induced bilayer permeabilization – OCer failed to active. To further examine possible Chol/StnII interactions, we measured Förster resonance energy transfer (FRET) between Trp in StnII and cholestatrienol (CTL), a fluorescent analog of Chol. We could show higher FRET efficiency between CTL and Trp:s in position 100 and 114 of StnII, when compared to three other Trp positions further away from the bilayer binding region of StnII. Taken together, our results suggest that StnII was able to attract Chol to its vicinity, maybe by showing affinity for Chol. SM interactions are known to be important for StnII binding to bilayers, and Chol is known to facilitate subsequent permeabilization of the bilayers by StnII. Our results help to better understand the role of these important membrane lipids for the bilayer properties of StnII.