Purified EDEM3 or EDEM1 alone produces determinant oligosaccharide structures from M8B in mammalian glycoprotein ERAD

Sequential mannose trimming of N-glycan, from M9 to M8B and then to oligosaccharides exposing the alpha 1,6-linked mannosyl residue (M7A, M6, and M5), facilitates endoplasmic reticulum-associated degradation of misfolded glycoproteins (gpERAD). We previously showed that EDEM2 stably disulfide-bonded to the thioredoxin domain-containing protein TXNDC11 is responsible for the first step (George et al., 2020). Here, we show that EDEM3 and EDEM1 are responsible for the second step. Incubation of pyridylamine-labeled M8B with purified EDEM3 alone produced M7 (M7A and M7C), M6, and M5. EDEM1 showed a similar tendency, although much lower amounts of M6 and M5 were produced. Thus, EDEM3 is a major alpha 1,2-mannosidase for the second step from M8B. Both EDEM3 and EDEM1 trimmed M8B from a glycoprotein efficiently. Our confirmation of the Golgi localization of MAN1B indicates that no other alpha 1,2-mannosidase is required for gpERAD. Accordingly, we have established the entire route of oligosaccharide processing and the enzymes responsible.

Automated summary

The sequential mannose trimming process in N-glycan modification involves EDEM2, EDEM3, and EDEM1 at different stages, with each enzyme playing a specific role in the endoplasmic reticulum-associated degradation of misfolded glycoproteins. The study establishes the entire route of oligosaccharide processing and confirms the Golgi localization of MAN1B, highlighting the essential enzymes involved in glycoprotein ERAD.