Engineering glycosyltransferases into glycan binding proteins using a novel mammalian surface display platform DOI
Sriram Neelamegham,

Ryoma Hombu,

Lauren Beatty

et al.

Research Square (Research Square), Journal Year: 2025, Volume and Issue: unknown

Published: May 2, 2025

Abstract Traditional lectins that bind cell-surface carbohydrate epitopes exhibit broad binding specificity. We tested the hypothesis more specific glycan-binding proteins may be developed by protein engineering of mammalian glycosyltransferases. Using rational design, we observed introducing an H302A mutation into porcine ST3Gal1, while abolishing enzymatic activity, resulted in a lectin displaying to sialylated core-2 O-linked glycans (i.e. Neu5Acα2-3Galβ1–3[GlcNAc(β1–6)]GalNAcα epitope). To expand on this, novel display platform screen for additional variants. One ST3Gal1 mutant (called ‘sCore2’) containing three mutations H302A/A312I/F313S exhibited improved properties. Spectral flow cytometry analysis with sCore2 shows individual blood cell types unique O-glycosylation profiles. Tissue microarray also reveals cell/tissue staining patterns sCore2. Overall, glycosyltransferases can engineered yield glycan proteins, suggesting similar approach extended other glycoenzymes also.

Language: Английский

Identification and Validation of Cyclic Peptides with Mucin-Selective, Location-Specific Binding in the Gastrointestinal Tract DOI Creative Commons
Deepak A. Subramanian, Albert Chin, Yunhua Shi

et al.

ACS Nano, Journal Year: 2025, Volume and Issue: unknown

Published: April 11, 2025

Oral drug delivery is a widely preferred method of administration due to its ease use and convenience for patients. Localization release in the gastrointestinal (GI) tract important treat localized diseases maximize absorption. However, achieving localization dynamic GI challenging. To address this challenge, we leveraged geographic diversity by targeting mucus layers, which coat epithelial surfaces. These composed mucin glycoproteins, are synthesized with unique chemical compositions expressed different regions, making them ideal targets localization. In article, identify cyclic peptides that bind selectively MUC2 (in intestines) MUC5AC stomach), serving as ligands these regions tract. We demonstrate effectiveness through vitro, ex vivo, vivo experiments, showing incorporating can increase binding selectivity 2-fold desired thus potentially overcoming challenges localizing distribution oral delivery. results indicate be used localize cargoes at certain sites body compared free drugs.

Language: Английский

Citations

0
Engineering glycosyltransferases into glycan binding proteins using a novel mammalian surface display platform DOI
Sriram Neelamegham,

Ryoma Hombu,

Lauren Beatty

et al.

Research Square (Research Square), Journal Year: 2025, Volume and Issue: unknown

Published: May 2, 2025

Abstract Traditional lectins that bind cell-surface carbohydrate epitopes exhibit broad binding specificity. We tested the hypothesis more specific glycan-binding proteins may be developed by protein engineering of mammalian glycosyltransferases. Using rational design, we observed introducing an H302A mutation into porcine ST3Gal1, while abolishing enzymatic activity, resulted in a lectin displaying to sialylated core-2 O-linked glycans (i.e. Neu5Acα2-3Galβ1–3[GlcNAc(β1–6)]GalNAcα epitope). To expand on this, novel display platform screen for additional variants. One ST3Gal1 mutant (called ‘sCore2’) containing three mutations H302A/A312I/F313S exhibited improved properties. Spectral flow cytometry analysis with sCore2 shows individual blood cell types unique O-glycosylation profiles. Tissue microarray also reveals cell/tissue staining patterns sCore2. Overall, glycosyltransferases can engineered yield glycan proteins, suggesting similar approach extended other glycoenzymes also.

Language: Английский

Citations

0