Study of the binding of viral glycoproteins by atomic force microscopy : application to Ebola and SARS-CoV-2
(2023)
Files
Thesis-QingrongZhang.pdf
Open Access - Adobe PDF
- 10.92 MB
Details
- Authors
- Supervisors
- Alsteens, David
- Abstract
- A multitude of human diseases stem from infections caused by enveloped viruses. For enveloped viruses, the infection begins with virions attaching to the surface of host cells, and then sets off a fusion process between viral and cellular membranes, enabling the viruses to penetrate the host cells. Grasping the molecular intricacies of how enveloped viruses enter cells is of utmost importance for the advancement of novel vaccines and therapeutics targeted at this process. Despite the wealth of insights provided by experimental studies into the entry of enveloped viruses, the finer details of their molecular mechanisms, particularly their dynamic aspects, remain significantly underexplored. This thesis centers around unraveling the kinetics and thermodynamics involved in the process of enveloped viruses entering cells. The approach taken involves utilizing atomic force microscopy (AFM)-based single molecule force spectroscopy (SMFS) to investigate the interaction between viruses and receptors at an individual virus level. Notably, a specialized AFM-SMFS operating in height-clamp mode was developed to dissect the binding of fusion peptides to the lipid membrane. We initially studied Ebola virus (EBOV) binding to vital receptors: the C-type lectin dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin receptor (DC-SIGNR) and the T-cell Ig and mucin domain 1 (TIM-1). We demonstrated that EBOV, activated by host enzymes, exhibits a preference for binding to TIM-1 over DC-SIGNR. TIM-1, binding to glycoprotein or phosphatidylserine in the envelope, functions as a dual-receptor for EBOV. Through analysis of kinetics and thermodynamics at the single virus level, we confirmed EBOV's strong affinity for both receptors. Our subsequent focus was on the interaction between the fusion peptide of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2 FP) and the host membrane. We demonstrated that cholesterol within the membrane, as opposed to sphingomyelin, increases the affinity of SARS-CoV-2 FP binding to the lipid membrane. Notably, the internal disulfide bond of SARS-CoV-2 FP was found to play a crucial role in enhancing its binding to the membrane—a conclusion supported by single-molecule kinetics and thermodynamics analyses. Additionally, we observed effective inhibition of SARS-CoV-2 infection through cholesterol removal from target cell membranes. In summary, this thesis uncovers new insights into enveloped virus entry mechanisms, spanning different steps and viruses. Additionally, the AFM experiments and theories introduced could have broader applications in studying novel virus-cell interactions and protein-membrane processes.
- Affiliations
Citations
Zhang, Q. (2023). Study of the binding of viral glycoproteins by atomic force microscopy : application to Ebola and SARS-CoV-2. https://hdl.handle.net/2078.5/235393