Journal Article

The 3-O-sulfation of heparan sulfate modulates protein binding and lyase degradation

Wolfert, Margreet A., Chopra, Pradeep, Joshi, Apoorva, Wu, Jiandong, Lu, Weigang, Yadavalli, Tejabhiram, Shukla, Deepak, Zaia, Joseph, Boons, Geert-Jan

2021

Overview

Humans express seven heparan sulfate (HS) 3-O-sulfotransferases that differ in substrate specificity and tissue expression. Although genetic studies have indicated that 3-O-sulfated HS modulates many biological processes, ligand requirements for proteins engaging with HS modified by 3-O-sulfate (3-OS) have been difficult to determine. In particular, the context in which the 3-OS group needs to be presented for binding is largely unknown. We describe herein a modular synthetic approach that can provide structurally diverse HS oligosaccharides with and without 3-OS. The methodology was employed to prepare 27 hexasaccharides that were printed as a glycan microarray to examine ligand requirements of a wide range of HS-binding proteins. The binding selectivity of antithrombin-III (AT-III) compared well with anti-Factor Xa activity supporting robustness of the array technology. Many of the other examined HS-binding proteins required an IdoA2S-GlcNS3S6S sequon for binding but exhibited variable dependence for the 2-OS and 6-OS moieties, and a GlcA or IdoA2S residue neighboring the central GlcNS3S. The HS oligosaccharides were also examined as inhibitors of cell entry by herpes simplex virus type 1, which, surprisingly, showed a lack of dependence of 3-OS, indicating that, instead of glycoprotein D (gD), they competitively bind to gB and gC. The compounds were also used to examine substrate specificities of heparin lyases, which are enzymes used for depolymerization of HS/heparin for sequence determination and production of therapeutic heparins. It was found that cleavage by lyase II is influenced by 3-OS, while digestion by lyase I is only affected by 2-OS. Lyase III exhibited sensitivity to both 3-OS and 2-OS.

• Publisher: National Academy of Sciences
• Subject: Acetylglucosamine - chemistry / Acetylglucosamine - metabolism / Antithrombin III - chemistry / Antithrombin III - genetics / Antithrombin III - metabolism / Binding Sites / Binding, Competitive / Carbohydrate Sequence / Cell Line / Chemistry / Cornea - cytology / Cornea - metabolism / Epithelial Cells - metabolism / Epithelial Cells - pathology / Epithelial Cells - virology / Factor Xa - chemistry / Factor Xa - genetics / Factor Xa - metabolism / Factor Xa Inhibitors - chemistry / Factor Xa Inhibitors - metabolism / Gene Expression / Glucuronic Acid - chemistry / Glucuronic Acid - metabolism / Heparin Lyase - chemistry / Heparin Lyase - genetics / Heparin Lyase - metabolism / Heparitin Sulfate - chemistry / Heparitin Sulfate - metabolism / Herpesvirus 1, Human - growth & development / Herpesvirus 1, Human - metabolism / Host-Pathogen Interactions - genetics / Humans / Isoenzymes - chemistry / Isoenzymes - genetics / Isoenzymes - metabolism / Microarray Analysis / Physical Sciences / Protein Binding / Proteolysis / Small Molecule Libraries / Substrate Specificity / Sulfates - chemistry / Sulfates - metabolism / Sulfotransferases - chemistry / Sulfotransferases - genetics / Sulfotransferases - metabolism / Viral Envelope Proteins - chemistry / Viral Envelope Proteins - genetics / Viral Envelope Proteins - metabolism