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Updated: Jan 10, 2026

Targeting Cysteine Thiols for in Vitro Site-specific Glycosylation of Recombinant Proteins
Published on: October 4, 2017
Structural basis of regulated N-glycosylation at the secretory translocon
Melvin Yamsek1, Mengxiao Ma2,3, Roshan Jha1
1Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL, USA.
Abstract:
Most human secretory pathway proteins are N-glycosylated by oligosaccharyltransferase (OST) complexes as they enter the endoplasmic reticulum (ER)1-3. Recent work revealed a substrate-assisted mechanism by which N-glycosylation of the chaperone glucose-regulated protein 94 (GRP94) is regulated to control cell surface receptor signalling4. Here we report the structure of a natively isolated GRP94 folding intermediate tethered to a specialized CCDC134-bound translocon. Together with functional analysis, the data reveal how a conserved N-terminal extension in GRP94 inhibits OST-A and how structural rearrangements within the translocon shield the tethered nascent chain from inappropriate OST-B glycosylation. These interactions depend on a hydrophobic CCDC134 groove, which recognizes a non-native conformation of nascent GRP94. Our results define a mechanism of regulated N-glycosylation and illustrate how the nascent chain remodels the translocon to facilitate its own biogenesis.
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