Parallel Glyco-SPOT Synthesis of Glycopeptide Libraries.

Journal Article (Journal Article)

Glycan recognition is typically studied using free glycans, but glycopeptide presentations represent more physiological conditions for glycoproteins. To facilitate studies of glycopeptide recognition, we developed Glyco-SPOT synthesis, which enables the parallel production of diverse glycopeptide libraries at microgram scales. The method uses a closed system for prolonged reactions required for coupling Fmoc-protected glycoamino acids, including O-, N-, and S-linked glycosides, and release conditions to prevent side reactions. To optimize reaction conditions and sample reaction progress, we devised a biopsy testing method. We demonstrate the efficient utilization of such microscale glycopeptide libraries to determine the specificity of glycan-recognizing antibodies (e.g., CTD110.6) using microarrays, enzyme specificity on-array and in-solution (e.g., ST6GalNAc1, GCNT1, and T-synthase), and binding kinetics using fluorescence polarization. We demonstrated that the glycosylation on these peptides can be expanded using glycosyltransferases both in-solution and on-array. This technology will promote the discovery of biological functions of peptide modifications by glycans.

Full Text

Duke Authors

Cited Authors

  • Mehta, AY; Veeraiah, RKH; Dutta, S; Goth, CK; Hanes, MS; Gao, C; Stavenhagen, K; Kardish, R; Matsumoto, Y; Heimburg-Molinaro, J; Boyce, M; Pohl, NLB; Cummings, RD

Published Date

  • September 17, 2020

Published In

Volume / Issue

  • 27 / 9

Start / End Page

  • 1207 - 1219.e9

PubMed ID

  • 32610041

Pubmed Central ID

  • PMC7556346

Electronic International Standard Serial Number (EISSN)

  • 2451-9448

Digital Object Identifier (DOI)

  • 10.1016/j.chembiol.2020.06.007

Language

  • eng

Conference Location

  • United States