Cell surfaces are glycosylated in various ways with high heterogeneity, which usually leads to ambiguous conclusions about glycan-involved biological functions. Here, we describe a two-step chemoenzymatic approach for N-glycan-subtype-selective editing on the surface of living cells that consists of a first ‘delete’ step to remove heterogeneous N-glycoforms of a certain subclass and a second ‘insert’ step to assemble a well-defined N-glycan back onto the pretreated glyco-sites. Such glyco-edited cells, carrying more homogeneous oligosaccharide structures, could enable precise understanding of carbohydrate-mediated functions. In particular, N-glycan-subtype-selective remodeling and imaging with different monosaccharide motifs at the non-reducing end were successfully achieved. Using a combination of the expression system of the Lec4 CHO cell line and this two-step glycan-editing approach, opioid receptor delta 1 (OPRD1) was investigated to correlate its glycostructures with the biological functions of receptor dimerization, agonist-induced signaling and internalization.
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All of the data are available in the paper and its Supplementary Information. Synthetic procedures and the characterization data of the new N-glycan substrates are also provided. Supporting data and processing protocols of cell imaging, FRET, FACS and western blot gels are available in the Supplementary files. The sequence data for the OPRD1 plasmid have been deposited to GenBank (accession code MN922300).
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This work was supported by the National Natural Science Foundation of China (no. 21877116, to W.H.), the National Science & Technology Major Project ‘Key New Drug Creation and Manufacturing Program’ of China (no. 2018ZX09711002-006, to W.H.) and the China Postdoctoral Science Foundation (no. BX20180339, to F.T., and no. 2018M642120, to F.T.). We thank R. Huang’s colleagues for their kind help with imaging studies. We also thank P. Wu of the Scripps Research Institute for providing us with Lec4 CHO cells.
The authors declare no competing interests.
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Tang, F., Zhou, M., Qin, K. et al. Selective N-glycan editing on living cell surfaces to probe glycoconjugate function. Nat Chem Biol 16, 766–775 (2020). https://doi.org/10.1038/s41589-020-0551-8
Nature Methods (2020)