Structural insights into TRAP association with ribosome-Sec61 complex and translocon inhibition by a CADA derivative

During cotranslational translocation, the signal peptide of a nascent chain binds Sec61 translocon to initiate protein transport through the endoplasmic reticulum (ER) membrane. Our cryo-electron microscopy structure of ribosome-Sec61 shows binding of an ordered heterotetrameric translocon-associated protein (TRAP) complex, in which TRAP-gamma is anchored at two adjacent positions of 28S ribosomal RNA and interacts with ribo-somal protein L38 and Sec61 alpha/gamma. Four transmembrane helices (TMHs) of TRAP-gamma cluster with one C-terminal helix of each alpha, 13, and delta subunits. The seven TMH bundle helps position a crescent-shaped trimeric TRAP-alpha/ 13/delta core in the ER lumen, facing the Sec61 channel. Further, our in vitro assay establishes the cyclotriazadisul-fonamide derivative CK147 as a translocon inhibitor. A structure of ribosome-Sec61-CK147 reveals CK147 binding the channel and interacting with the plug helix from the lumenal side. The CK147 resistance mutations surround the inhibitor. These structures help in understanding the TRAP functions and provide a new Sec61 site for designing translocon inhibitors.

Automated summary

During cotranslational translocation, the signal peptide of a nascent chain binds Sec61 translocon to initiate protein transport through the endoplasmic reticulum (ER) membrane. Our cryo-electron microscopy structure of ribosome-Sec61 shows binding of an ordered heterotetrameric translocon-associated protein (TRAP) complex, in which TRAP-gamma is anchored at two adjacent positions of 28S ribosomal RNA and interacts with ribosomal protein L38 and Sec61 alpha/gamma. Four transmembrane helices (TMHs) of TRAP-gamma cluster with one C-terminal helix of each alpha, 13, and delta subunits.