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bioRxiv · 10.1101/2025.08.21.671495

Identification and characterization of a novel exo-glycosidase activity of NGLY1 on ENGase-digested N-GlcNAc proteins in vitro

Ye, Y.·Lu, X.·Zhang, Y.·Lin, Q.·Qian, X.·Lin, J.·Yu, Z.·Lyu, S.·Sun, G.·CHEN, L.
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Abstract

N-Glycanase 1 (NGLY1) is known for its activity to completely remove conjugated N-glycans from glycoproteins. Early studies illustrated that NGLY1 defects were associated with a rare disease named NGLY1-related congenital disorder of deglycosylation (NGLY1-CDDG). Although extensive research has been conducted over the past decade on the biological impact of NGLY1s endo-(N-)glycanase activity in cells and on disease, whether NGLY1 also exhibits an exo-(N-)glycanase activity remains open. In this study, an exo-(N-)glycanase activity of NGLY1 on the N-GlcNAc proteins was firstly reported and characterized in vitro. To distinguish for the conventional endo-(N-)glycanase activity and the newly reported exo-(N-)glycanase activity of NGLY1, the active sites of exo-NGLY1 were predicted and compared with those of endo-NGLY1. In addition, the correlation between NGLY1s exo-(N-)glycanase activity in vitro and NGLY1-CDDGs clinical characterization was investigated and discussed. The observation of NGLY1s novel exo-(N-)glycanase activity expanded the functional repertoire of NGLY1 beyond canonical endo-(N-)glycanase-mediated deglycosylation, and may provide a new framework to explain the clinical heterogeneity of NGLY1-CDDG.

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Ye, Y., Lu, X., Zhang, Y., Lin, Q., Qian, X., Lin, J., Yu, Z., Lyu, S., Sun, G., CHEN, L.. 2025-08-21. Identification and characterization of a novel exo-glycosidase activity of NGLY1 on ENGase-digested N-GlcNAc proteins in vitro. https://doi.org/10.1101/2025.08.21.671495

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