PXD072144

PXD072144 is an original dataset announced via ProteomeXchange.

Title	Functional assays of human ALG9
Description	Asparagine-linked glycans are essential for the maturation and function of most eukaryotic secretory proteins. Biosynthesis and transfer of the GlcNAc2Man9Glc3 glycan occurs on the endoplasmic reticulum (ER) membrane, in a highly conserved process involving more than a dozen membrane proteins whose dysfunction is linked to severe congenital disorders of glycosylation (CDGs). Three membrane-integral mannosyltransferases ALG3, ALG9 and ALG12 mediate four consecutive mannosylation reactions that convert GlcNAc2Man5 to GlcNAc2Man9. Using chemo-enzymatically synthesised lipid-linked glycan donor and acceptor analogues, we recapitulated this biosynthetic pathway in vitro. High-resolution cryo-EM structures of pseudo-Michaelis complexes of each step revealed the molecular basis of how the branched glycan is accurately synthesised in sequential reactions. Molecular dynamics simulations, mutagenesis, and functional assays uncovered a subtle but high precision mechanisms selecting the dolichylphosphomannose donor substrate over the also present epimer, dolichylphosphoglucose. Together, our results provide mechanistic explanations for enzyme disfunction in CDG and offer opportunities for N-glycan engineering.
HostingRepository	PRIDE
AnnounceDate	2026-05-30
AnnouncementXML	Submission_2026-05-30_05:57:40.094.xml
DigitalObjectIdentifier	https://doi.org/10.6019/PXD072144
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	J. Andrew Alexander
SpeciesList	scientific name: synthetic construct; NCBI TaxID: NEWT:32630;
ModificationList	amidated residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2025-12-18 06:56:13	ID requested
⏵ 1	2026-05-30 05:57:40	announced

10.1038/S41589-026-02164-7;

10.6019/PXD072144;

submitter keyword: ALG9, oligomannose, N-glycans

Kaspar P. Locher
contact affiliation	Institute of Molecular Biology and Biophysics, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland
contact email	locher@mol.biol.ethz.ch
lab head
J. Andrew Alexander
contact affiliation	ETH Zürich
contact email	andrew.alexander@mol.biol.ethz.ch
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD072144

PRIDE project URI

• PRIDE
  1. PXD072144
     1. Label: PRIDE project
     2. Name: Functional assays of human ALG9