A Glycoengineered Enzyme with Multiple Mannose-6-Phosphates Is Internalized into Diseased Cells to Restore Its Activity in Lysosomes

Cell Chemical Biology
Ji Young HyunInjae Shin

Abstract

In this study we developed an efficient method to prepare glycoengineered β-N-acetylhexosaminidase containing multiple mannose-6-phosphates (M6Ps) by combining genetic code expansion with bioorthogonal ligation techniques. We found that multiple M6P-conjugated enzymes were produced with a high efficiency by using combined techniques. Importantly, glycoengineered enzymes entered lysosomes of patient-derived primary cells, which lack endogenous lysosomal β-N-acetylhexosaminidase, more readily than commercialized human β-hexosaminidase. Moreover, glycoengineered enzymes successfully removed GM2-ganglioside stored in lysosomes of diseased cells, indicating that its activity is restored in diseased cells. We also synthesized and applied a lysosome-targeting fluorogenic substrate to monitor endogenous and supplemental glycoengineered β-N-acetylhexosaminidase activities in lysosomes. The results of this study indicate that the present strategy, which relies on genetic code expansion and bioorthogonal ligation techniques, is highly attractive to generate multi-M6P-containing lysosomal enzymes that can be used to study lysosomal storage disorders associated with lysosomal enzyme deficiencies.

Citations

Aug 10, 2019·Human Gene Therapy·Valentina Poletti, Alessandra Biffi
May 5, 2020·Glycobiology·Krzysztof MikolajczykMarcin Czerwinski
Feb 13, 2019·Chemical Science·Sang-Hyun ParkInjae Shin
Aug 3, 2019·European Journal of Medicinal Chemistry·Lucia BiasuttoMario Zoratti
May 3, 2021·Current Opinion in Chemical Biology·Nikki DellasGjalt W Huisman
Aug 5, 2021·Chemical Society Reviews·Yujun KimInjae Shin
Nov 25, 2020·ACS Chemical Biology·Ji Young HyunInjae Shin

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