Extended Data Fig. 2: Characterization of GABRA1 C-terminal TMD insertion.
From: EMC rectifies the topology of multipass membrane proteins
Domain diagram of 23L-GABRA1 is shown (top): a glycosylated N-tail is followed by a TMD comprised of 23 leucine residues (23L), a cytosolic loop (~100aa), and TMD4 of GABRA1 that is followed by a translocated and glycosylated C-tail. 35S-methionine labeled 23L-GABRA1 (middle) or ASGR1 (bottom) were translated in the presence of SPCs from WT, ΔEMC (ΔE), ΔTMCO1 (ΔT) or ΔEMCΔTMCO1 (ΔET) cells. Translated products were analyzed directly by SDS-PAGE and autoradiography. Where indicated, Sec61 inhibitor ApraA was included in the translation reaction. Substrates with different glycosylation states are indicated. C-tail translocation of 23L-GABRA1 was quantified by calculating the percentage of double glycosylated products among all glycosylated products.
