Biochemistry Department, Faculty of Science and Engineering, Åbo Akademi University, Turku, Finland.
Departamento de Bioquímica y Biología Molecular, Universidad Complutense, Madrid, Spain.
Sci Rep. 2022 Oct 15;12(1):17328. doi: 10.1038/s41598-022-21750-y.
Sticholysins are α-pore-forming toxins produced by the sea-anemone Stichodactyla helianthus. These toxins exert their activity by forming pores on sphingomyelin-containing membranes. Recognition of sphingomyelin by sticholysins is required to start the process of pore formation. Sphingomyelin recognition is coupled with membrane binding and followed by membrane penetration and oligomerization. Many features of these processes are known. However, the extent of contact with each of the different kinds of lipids present in the membrane has received little attention. To delve into this question, we have used a phosphatidylcholine analogue labeled at one of its acyl chains with a doxyl moiety, a known quencher of tryptophan emission. Here we present evidence for the contact of sticholysins with phosphatidylcholine lipids in the sticholysin oligomer, and for how each sticholysin isotoxin is affected differently by the inclusion of cholesterol in the membrane. Furthermore, using phosphatidylcholine analogs that were labeled at different positions of their structure (acyl chains and headgroup) in combination with a variety of sticholysin mutants, we also investigated the depth of the tryptophan residues of sticholysins in the bilayer. Our results indicate that the position of the tryptophan residues relative to the membrane normal is deeper when cholesterol is absent from the membrane.
海葵素是由海葵属海葵产生的α-孔形成毒素。这些毒素通过在含有神经鞘磷脂的膜上形成孔发挥作用。海葵素识别神经鞘磷脂是开始形成孔的过程所必需的。海葵素识别与膜结合耦联，随后是膜穿透和寡聚化。这些过程的许多特征是已知的。然而，对于膜中存在的每种不同类型的脂质的接触程度，关注甚少。为了深入研究这个问题，我们使用了一种在其酰基链之一上用 doxyl 部分标记的磷脂酰胆碱类似物，doxyl 部分是色氨酸发射的已知猝灭剂。在这里，我们提供了证据表明海葵素与磷脂酰胆碱脂质在海葵素寡聚体中的接触，以及胆固醇在膜中的存在如何不同地影响每种海葵素同工毒素。此外，使用在其结构的不同位置（酰基链和头基）处标记的磷脂酰胆碱类似物，并结合各种海葵素突变体，我们还研究了海葵素色氨酸残基在双层中的深度。我们的结果表明，当膜中没有胆固醇时，色氨酸残基相对于膜法线的位置更深。
