Glycosylation of Receptor Binding Domain of SARS-CoV-2 S-Protein Influences on Binding to Immobilized DNA Aptamers

Abstract

Nucleic acid aptamers specific to S-protein and its receptor binding domain (RBD) of SARS-CoV-2 (severe acute respiratory syndrome-related coronavirus 2) virions are of high interest as potential inhibitors of viral infection and recognizing elements in biosensors. Development of specific therapy and biosensors is complicated by an emergence of new viral strains bearing amino acid substitutions and probable differences in glycosylation sites. Here, we studied affinity of a set of aptamers to two Wuhan-type RBD of S-protein expressed in Chinese hamster ovary cell line and Pichia pastoris that differ in glycosylation patterns. The expression system for the RBD protein has significant effects, both on values of dissociation constants and relative efficacy of the aptamer binding. We propose glycosylation of the RBD as the main force for observed differences. Moreover, affinity of a several aptamers was affected by a site of biotinylation. Thus, the robustness of modified aptamers toward new virus variants should be carefully tested.

An engineered SARS-CoV-2 receptor-binding domain produced in Pichia pastoris as a candidate vaccine antigen

“…Unlike other reports expressing the RBD in P. pastoris [29] , [30] , [31] here the potential N-glycosylation site at N331 in C-RBD-H6 PP was included. Although inclusion of N331 leads to more heterogeneous N-glycosylation [3] , [32] , [33] , it should be noted that P. pastoris N-glycans are often hypermannosylated [34] , [35] , and mannosylation enhances the activation of antigen-presenting cells like macrophages and dendritic cells, increasing immunogenicity over that of non-glycosylated counterparts [8] , [36] , [37] , [38] . In fact, others [29] , [31] used a lipid-modified alum adjuvant or a saponin-based adjuvant, respectively, to overcome the poor immunogenicity of their N331-less RBD antigens.…”

Nanomaterials‐based sensors for the detection of COVID‐19: A review

“…13 Thus, in the past couple of years, a lot of work has been conducted on rapidly detecting COVID‐19 using novel biosensors. 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 Likewise, many prominent researchers have spent a lot of time compiling several review articles about recent COVID diagnosis and treatment using different sensing strategies. 7 , 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 However, to the best of our knowledge, there is no systematic review article that extensively sums up the recent developments and research on nanomaterial‐based biosensors for rapid, sensitive, and selective detection of the SARS‐CoV‐2.…”

“…The National Institutes of Health strategic plan for COVID‐19 research calls for the need “to improve the basic understanding of SARS‐CoV‐2 and COVID 19 and develop the necessary tools and approaches to diagnose, prevent, and treat this disease,” highlighting the impetus on the rapid and reliable diagnosis of the coronavirus 13 . Thus, in the past couple of years, a lot of work has been conducted on rapidly detecting COVID‐19 using novel biosensors 14–21 . Likewise, many prominent researchers have spent a lot of time compiling several review articles about recent COVID diagnosis and treatment using different sensing strategies 7,22–30 .…”

Comprehensive role of SARS‐CoV‐2 spike glycoprotein in regulating host signaling pathway

“…For example, elegant molecular dynamics simulations show that N‐glycans at sites N165 and N234 of S protein in modulating the conformational dynamics of the spike's RBD, which is responsible for ACE2 recognition, 77 while glycans at N090, N322, and N546 of ACE2 directly mediate the interaction with S trimer 75 . Aptamer studies also show that glycosylation of the RBD protein has significant effects on values of dissociation constants and the relative efficacy of the aptamer binding 78 . Thus, variations in glycan occupancy or processing at these sites could alter the affinity of the SARS‐CoV‐2–ACE2 interaction and modulate infectivity.…”

“… 75 Aptamer studies also show that glycosylation of the RBD protein has significant effects on values of dissociation constants and the relative efficacy of the aptamer binding. 78 Thus, variations in glycan occupancy or processing at these sites could alter the affinity of the SARS‐CoV‐2–ACE2 interaction and modulate infectivity. Furthermore, S protein can interact with SARS‐CoV‐2 attachment receptor type C lectin through a high‐mannose N‐glycan structure, and this interaction can be blocked by glycosylation inhibitors.…”