Characterization of glycan-protein interactions in SARS-CoV-2 and Parkinson's disease

Gelbach, Adrianne L
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Linhardt, Robert J.
Gilbert, Susan P.
Eliezer, David
Wang, Chunyu
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ABSTRACT. Heparan sulfate (HS) acts as a co-receptor of angiotensin-converting enzyme 2 (ACE2) by interacting with severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) spike glycoprotein (SGP), facilitating host cell entry of SARS-CoV-2 virus. Heparin, a highly sulfated version of heparan sulfate (HS), interacts with a variety of proteins playing key roles in many physiological and pathological processes. In this study, SARS-CoV-2 SGP receptor binding domain (RBD) of wild type (WT), Delta and Omicron variants were expressed in Expi293F cells and used in the kinetic and structural analysis on their interactions with heparin. Surface plasmon resonance (SPR) analysis showed the binding kinetics of SGP RBD from WT and Delta variants were very similar while Omicron variant SGP showed a much higher association rate. The SGP from Delta and Omicron showed higher affinity (KD) to heparin than the WT SGP. Competition SPR studies using heparin oligosaccharides indicated that optimal binding of SGP RBDs to heparin requires chain length greater than 18. Chemically modified heparin derivatives all showed reduced interactions in competition assays suggesting that all the sulfo groups in the heparin polysaccharide were critical for binding SGP RBDs with heparin. These interactions with heparin are pH sensitive. Acidic pH (pH 6.5, 5.5, 4.5) greatly increased the binding of WT and Delta SGP RBDs to heparin, while acidic pH only slightly reduced the binding of Omicron SGP RBD to heparin compared to binding at pH 7.3. In contrast, basic pH (pH 8.5) greatly reduced the binding of Omicron SGP RBDs to heparin, with much less effects on WT or Delta. The pH dependence indicates different charged residues were present at the Omicron SGP-heparin interface. Detailed kinetic and structural analysis of the interactions of SARS-CoV-2 SGP RBDs with heparin provides important information for designing anti-SARS-CoV-2 molecules.Heparan sulfate on neuronal surface also plays important roles in the prion-like spread of -synuclein (aS) pathology in Parkinson’s disease (PD). Using similar SPR methods employed in heparin-SPG studies, we show that aS binds heparin with 0.4 M affinity, and that N-sulfation is the most important sulfation pattern for heparin-aS interaction. In addition, glycan chain length plays a crucial role in heparin-aS interaction, with a minimal chain length of 16 saccharide units required for optimal interaction. NMR has been used to characterize the binding sites in aS. Based on chemical shift perturbation and peak intensity change, C-terminal acidic tail is minimally involved while the non amyloid component (NAC) domain is the most important region for heparin binding. K81, K59 and K61, within the imperfect KTKEGV repeats, likely provide the crucial positive charges for interaction with heparin. Detailed SPR and NMR provide novel insights towards molecular mechanisms and therapeutic intervention of PD.
August 2022
School of Science
Dept. of Biological Sciences
Rensselaer Polytechnic Institute, Troy, NY
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