Datasets from publication: Front. Catal.Sec. BiocatalysisVolume 3 - 2023 | doi: 10.3389/fctls.2023.1275281

Introduction: Glycopeptides contain carbohydrate moieties (glycans) covalently attached to the side chain and/or terminal peptide units. Since glycans are present on cell surfaces, these constructs can potentially address a wide array of therapeutic functions. To overcome the deficiencies associated with current synthetic routes to glycopeptides, such as costly processes and toxic reagents, this work aimed to develop versatile environmentally friendlyprotease-catalyzed peptide synthetic routes to peptides decorated with aglycan at their N-terminus.Methods: “Grafters” were first synthesized that consist of a glycanconjugated directly, or through a spacer, to the amine group of L-Phe-ethylester (Phe-OEt). The role of Phe-OEt is to increase the conjugate’s recognitionby the protease (papain) catalytic active site. A series of grafters weresynthesized with variation of the glycan structure, linkage-chemistry, andpresence of an oligo (ethylene glycol) “spacer” of varied length between theglycan and Phe-OEt moiety. High grafter efficiency will result by the successfulacceptance of the grafter at the enzymes S1/S2 subsites, formation of an acyl enzymecomplex and subsequent conversion to glycan-terminated oligo(Leu)x (x ≥ 1), as opposed to construction of non-glycan N-terminated oligo(Leu)x. Results and discussion: While glycan-Phe-OEt grafters without a spacer between theglycan and Phe-OEt resulted in low grafter efficiency (8.3% ± 2.0%),insertion of a short oligo(ethylene glycol) spacer between the glycan andPhe-OEt moieties (glycan-PEGn-Phe-OEt, n ≥ 3) increased the grafterefficiency by 3-fold–24.5% ± 1.8%. In addition, computational modeling wasperformed using Rosetta software provided insights on a molecular level ofhow grafter efficiency is influenced by the PEG spacer length.