Structure of the Inhibited State of the Sec Translocon

Abstract

Summary Protein secretion in eukaryotes and prokaryotes involves a universally conserved protein translocation channel formed by the Sec61 complex. Unrelated small-molecule natural products and synthetic compounds inhibit Sec61 with differential effects for different substrates or for Sec61 from different organisms, making this a promising target for therapeutic intervention. To understand the mode of inhibition and provide insight into the molecular mechanism of this dynamic translocon, we determined the structure of mammalian Sec61 inhibited by the Mycobacterium ulcerans exotoxin mycolactone via electron cryo-microscopy. Unexpectedly, the conformation of inhibited Sec61 is optimal for substrate engagement, with mycolactone wedging open the cytosolic side of the lateral gate. The inability of mycolactone-inhibited Sec61 to effectively transport substrate proteins implies that signal peptides and transmembrane domains pass through the site occupied by mycolactone. This provides a foundation for understanding the molecular mechanism of Sec61 inhibitors and reveals novel features of translocon function and dynamics.

Ipomoeassin-F inhibits the in vitro biogenesis of the SARS-CoV-2 spike protein and its host cell membrane receptor

“…Consistent with this unusual membrane topology, we found no indication that the membrane insertion of any of our OPG2-tagged ORF6 variants was reduced by Ipom-F, strongly suggesting that its association with the inner leaflet of the ER membrane does not require protein translocation via the central channel of the Sec61 translocon ( Gérard et al, 2020 ). We noted a subset of OPG2–ORF6–OPG2 species bearing only a single N-glycan was also clearly present in the membrane-associated fraction with or without Ipom-F treatment ( Fig.…”

Ipomoeassin-F inhibits thein vitrobiogenesis of the SARS-CoV-2 spike protein and its host cell membrane receptor

“…It seems likely that viral type II TMPs will also be inhibited by Ipom-F (cf. Morel et al, 2018; Zong et al, 2019; Gérard et al, 2020) but, since SARS-CoV-2 has no such proteins, we have not addressed this issue.…”

Immunity against Mycobacterium ulcerans: The subversive role of mycolactone

“…94 Interestingly, the mutations conferring resistance to mycolactone clustered in the same region, 73 suggesting that structurally different Sec61α inhibitors share a common binding site and mode of action. Gérard et al 95 recently reported an electron cryo-microscopy structure of mycolactone bound Sec61α. Surprisingly, the mycolactone-binding pocket did not overlap with the region identified by resistance mutations, suggesting that such mutations may induce conformational changes indirectly preventing mycolactone binding.…”