9f5h

Publication Abstract from PubMed

Correct elaboration of N-linked glycans in the secretory pathway of human cells is essential in physiology. Early N-glycan biosynthesis follows an assembly line principle before undergoing crucial elaboration points that feature the sequential incorporation of the sugar N-acetylglucosamine (GlcNAc). The activity of GlcNAc transferase V (MGAT5) primes the biosynthesis of an N-glycan antenna that is heavily upregulated in cancer. Still, the functional relevance and substrate choice of MGAT5 are ill-defined. Here, we employ protein engineering to develop a bioorthogonal substrate analog for the activity of MGAT5. Chemoenzymatic synthesis is used to produce a collection of nucleotide-sugar analogs with bulky, bioorthogonal acylamide side chains. We find that WT-MGAT5 displays considerable activity toward such substrate analogues. Protein engineering yields an MGAT5 variant that loses activity against the native nucleotide sugar and increases activity toward a 4-azidobutyramide-containing substrate analogue. By such restriction of substrate specificity, we show that the orthogonal enzyme-substrate pair is suitable to bioorthogonally tag glycoproteins. Through X-ray crystallography and molecular dynamics simulations, we establish the structural basis of MGAT5 engineering, informing the design rules for bioorthogonal precision chemical tools.

A Bioorthogonal Precision Tool for Human N-Acetylglucosaminyltransferase V.,Liu Y, Bineva-Todd G, Meek RW, Mazo L, Piniello B, Moroz O, Burnap SA, Begum N, Ohara A, Roustan C, Tomita S, Kjaer S, Polizzi K, Struwe WB, Rovira C, Davies GJ, Schumann B J Am Chem Soc. 2024 Oct 2;146(39):26707-26718. doi: 10.1021/jacs.4c05955. Epub , 2024 Sep 17. PMID:39287665^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.