The Ca²⁺‐Regulated Photoprotein Obelin as a Tool for SELEX Monitoring and DNA Aptamer Affinity Evaluation

Abstract

Bioluminescent solid‐phase analysis was proposed to monitor the selection process and to determine binding characteristics of the aptamer–target complexes during design and development of the specific aptamers. The assay involves Ca2+‐regulated photoprotein obelin as a simple, sensitive and fast reporter. Applicability and the prospects of the approach were exemplified by identification of DNA aptamers to cardiac troponin I, a highly specific early biomarker for acute myocardial infarction. Two structurally different aptamers specific to various epitopes of troponin I were obtained and then tested in a model bioluminescent assay.

Enzymatic Responses to Low-Intensity Radiation of Tritium

“…Additionally, one of the simplest enzymatic bioassay systems, i.e., a Coelenteramide-containing Fluorescent Protein (CLM-FP), was used to evaluate the activation bioeffect of tritium. CLM-FPs are products of bioluminescent reactions of coelenterates that constitute a potential use in biomedicine as biomarkers [36][37][38]. The fluorescence characteristics of CLM-FPs were intensively studied [39][40][41][42][43][44][45][46]; they were found to be sensitive to external exposures: excitation energy [44], chemical agents [47,48], and radioactivity [49,50].…”

“…Our studies of enzymatic responses can provide a more detailed understanding of the mechanisms of low-dose effects and expand the application of enzyme systems for the diagnosis and/or regulation of intracellular processes. For example, bioluminescent reactions of coelenterates are widely used as genetically encoded markers in biological and medical research [36][37][38], and the parallel use of the products of their reactions (CLM-FPs) to assess the radiation-induced processes in the intracellular media would provide these reagents with multifunctionality.…”

Development of Nucleic‐Acid‐Based Electrochemical Biosensors for Clinical Applications

“…To take advantage of the better storage stability and shelf life of nucleic acids compared to antibodies, a range of aptamers have been selected for targeting cardiac biomarkers, including troponin, [207–211] myoglobin, [212, 213] creatine kinase, [214] heart‐type fatty‐acid‐binding protein (hFABP), [215] B‐type natriuretic peptide (BNP), [216–218] Interleukin‐6 (IL‐6), [219–222] and C‐reactive protein (CRP) [223–226] . Aptamer‐based biosensors have been developed for different cardiac biomarkers, where electrodes modified with target specific aptamers are used.…”

Coelenterazine-Dependent Luciferases as a Powerful Analytical Tool for Research and Biomedical Applications

“…Using obelin as a reporter—a convenient technique—was developed for fast and easy monitoring of the DNA library enrichment and evaluating the affinity and specificity of the individual aptamers during the SELEX procedure [ 81 ]. The approach was applied for the selection and development of 2′-F-Py RNA aptamers to general and glycated hemoglobins—biomarkers of diabetes mellitus [ 82 ] and DNA aptamers to the cardiac troponin I—a specific biomarker for acute myocardial infarction [ 81 ].…”