Selection of a DNA aptamer against norovirus capsid protein VP1

Abstract

The genetically and antigenically diverse group of noroviruses is the major cause of human viral epidemic gastroenteritis worldwide. Virus detection and control are thus crucial topics when aiming at containing and preventing the resulting large and often persisting outbreaks. Aptamers provide a promising alternative to antibodies concerning their ability to bind and thus detect and influence bio-active molecules. These small, single-stranded oligonucleotides are able to bind to a multitude of possible target molecules with high affinity. For a specific target the highest affinity aptamers are found by screening a randomized library. In this work a DNA aptamer capable of binding to the norovirus genotype II.4 capsid protein VP1 was found. The general approach is thereby not limited to norovirus capsid, but could be extended to almost any kind of biologically relevant molecule. The development of the library enrichment was further computationally analyzed in order to describe the enrichment during screening. This is the basis for a later extensive characterization of both target and aptamers that could lead to insights regarding the functional coherence of both partners. An abstract model describing this coherence could be utilized to generate a target-specific library, from which future aptamer screening runs could benefit.

Generation and characterization of nucleic acid aptamers targeting the capsid P domain of a human norovirus GII.4 strain

“…However, the work focused primarily on innovations in bioinformatic analysis and protein-aptamer modeling rather than the functional binding characterization reported here. Interestingly, the aptamers produced by Giamberardino et al (2013) and Beier et al (2014) had G values similar to M1 and M6-2, but the sequences and secondary structures differed from ours. In both papers, the aptamers produced were never applied for capture or detection of human NoV in outbreak-derived stool specimens, and aptamer binding to the intact capsid of only one genotype of human NoV was confirmed for any of the reported aptamers.…”

“…This aptamer was used as a recognition element in a voltammetry-based biosensor. Beier et al (2014) created DNA aptamers using an unspecified GII.4 strain's entire major capsid protein (VP1) by a different SELEX process than ours. However, the work focused primarily on innovations in bioinformatic analysis and protein-aptamer modeling rather than the functional binding characterization reported here.…”

Selection, Characterization and Application of Nucleic Acid Aptamers for the Capture and Detection of Human Norovirus Strains

“…Two recent papers report on the development of aptamers directed against noroviruses [23] [24]. Specifically, Giamberardino et al (2013) [23] produced a DNA aptamer against the cultivable surrogate murine norovirus (MNV) that displayed cross-reactivity to a GII.3 HuNoV VLP.…”

“…Common to our approach, these investigators directed the SELEX process against the whole virus and used a similar number of selection rounds. Beier et al (2014) [24] used a SELEX process directed against the capsid protein (VP1) of a GII.4 HuNoV to produce an aptamer for which they simulated the binding characteristics using predictive modeling. They did not evaluate the performance of the aptamer for virus capture or detection in clinical or food samples.…”

Human Norovirus Aptamer Exhibits High Degree of Target Conformation-Dependent Binding Similar to That of Receptors and Discriminates Particle Functionality

“…They offer the advantages of low cost; ease of synthesis, purification, and functional modification; and high stability. Multiple aptamers have been generated for human norovirus binding, some of which are broadly reactive (19–22). Evidence has been presented that aptamers tend to bind their targets with multiple different discriminatory interactions in unique ways (23), and as a consequence, instances have been reported in which aptamers did not bind denatured target proteins (24–26).…”