Small-molecule binding aptamers: Selection strategies, characterization, and applications

Aptamers are single-stranded, synthetic oligonucleotides that fold into 3-dimensional shapes capable of binding non-covalently with high affinity and specificity to a target molecule. They are generated via an in vitro process known as the Systematic Evolution of Ligands by EXponential enrichment, from which candidates are screened and characterized, and then used in various applications. These applications range from therapeutic uses to biosensors for target detection. Aptamers for small molecule targets such as toxins, antibiotics, molecular markers, drugs, and heavy metals will be the focus of this review. Their accurate detection is needed for the protection and wellbeing of humans and animals. However, the small molecular weights of these targets, including the drastic size difference between the target and the oligonucleotides, make it challenging to select, characterize, and apply aptamers for their detection. Thus, recent (since 2012) notable advances in small molecule aptamers, which have overcome some of these challenges, are presented here, while defining challenges that still exist are discussed

Effect of point mutations on the secondary structure and membrane interaction of antimicrobial peptide anoplin

Anoplin (GLLKRIKTLL-NH2) is the smallest linear α-helical antimicrobial peptide found naturally to date. Antibacterial and hemolytic properties of anoplin depend strongly on physicochemical properties. Two anoplin derivatives, anoplin-8K (Ano8K, GLLKTIKKLL-NH2) and anoplin-1K5 V8K (Ano1K5 V8K, KLLKVIKLL-NH2), were found to have increased bacterial and low hemolytic activity. In the present work physicochemical properties of these three peptides were studied by UV resonance Raman (UVRR) spectroscopy, Langmuir-Blodgett monolayer technique, and carboxyfluorescein (CF) leakage assay. UVRR data indicated that all three peptides adopt predominantly unordered conformation in aqueous buffer solution. In membrane-mimicking trifluoroethanol, the α-helical content increases for all three peptides with Ano1K5 V8K having the highest α-helix percentage, followed by Ano8K and anoplin. Critical micelle concentrations were found to be similar for all three peptides, and the saturation pressure decreases in the sequence Ano1K5 V8K, anoplin, Ano8K. Critical pressure of insertion was found to be greater for anionic lipid monolayer DPPG than for zwitterionic lipid DPPC indicating preferential adsorption of all three peptides to DPPG. Finally, membrane lytic activities of all three peptides toward various model lipid vesicles were compared through CF leakage assay. Overall the data indicate that antimicrobial activity of anoplin increases with charge, whereas membrane lytic activity correlates with peptides helicity and amphipathicity

Current status and future prospects for aptamer-based mycotoxin detection

Aptamers are single-stranded oligonucleotides with the ability to bind tightly and selectively to a target analyte. High-affinity and specific aptamers for a variety of mycotoxins have been reported over the past decade. Increasingly, these molecular recognition elements are finding applications in biosensors and assays for the detection of mycotoxins in a variety of complex matrixes. This review article highlights the mycotoxin aptamers that are available for mycotoxin detection and the array of biosensing platforms into which they have been incorporated. Key advantages that aptamers have over analogous technology, and areas in which these advantages may be applied for the benefit of practical mycotoxin detection, are also discussed

Analysis of In Vitro Aptamer Selection Parameters

Nucleic acid aptamers are novel molecular recognition tools that offer many advantages compared to their antibody and peptide-based counterparts. However, challenges associated with in vitro selection, characterization, and validation have limited their wide-spread use in the fields of diagnostics and therapeutics. Here, we extracted detailed information about aptamer selection experiments housed in the Aptamer Base, spanning over two decades, to perform the first parameter analysis of conditions used to identify and isolate aptamers de novo. We used information from 492 published SELEX experiments and studied the relationships between the nucleic acid library, target choice, selection methods, experimental conditions, and the affinity of the resulting aptamer candidates. Our findings highlight that the choice of target and selection template made the largest and most significant impact on the success of a de novo aptamer selection. Our results further emphasize the need for improved documentation and more thorough experimentation of SELEX criteria to determine their correlation with SELEX success