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Updated: Jan 9, 2026
Exploring Sequence Space to Identify Binding Sites for Regulatory RNA-Binding Proteins
Published on: August 9, 2019
Integrating Magnetic Bead-Based SELEX with In Silico Binding Analyses for the Identification of High-Affinity DNA
Darius R Martin1,2, Ayman Waddad3, Mohammed A Gafar4
1Department of Science, Technology and Innovation/TIA Nanotechnology Platform, Department of Biotechnology, Faculty of Natural Sciences, University of the Western Cape, Private Bag X17, Bellville, Cape Town 7535, South Africa.
Abstract:
This study aimed to identify aptamers that bind with high affinity to transgelin-2 (TAGLN2), a potential diagnostic biomarker for a number of diseases, such as cancer and tuberculosis, that is associated with cellular stress. Aptamers targeting recombinant TAGLN2 were selected through magnetic bead-based systematic evolution of ligands by exponential enrichment (SELEX). DNA sequence analysis and Geneious software analysis identified 10 unique aptamer sequences that potentially bind to TAGLN2. A phylogenetic analysis of these sequences shows that these sequences clustered into two clades based on sequence similarity, with the sequences in one of the clades (consisting of 4 unique sequences) showing higher similarity to each other. The secondary structures of aptamer sequences (aptamers 1, 2, 7, and 9) from this highly conserved clade were predicted using the M-fold web server. The binding of two of these aptamer sequences (aptamer 7 and 9) to recombinantly expressed TAGLN2 was verified by microscale thermophoresis (MST). The dissociation constants for aptamers 7 and 9 were determined to be 18 ± 1 and 50 ± 2 nM, respectively. In silico analysis was used to perform molecular docking and molecular dynamics (MD) simulations between TAGLN2 and all 4 aptamers. These in silico analyses support the finding that aptamers 7 and 9 have a high affinity for TAGLN2 and that aptamer 7 has a higher binding affinity than aptamer 9. However, based on the in silico analysis, aptamer 1 may be a stronger binder than aptamer 7. This study demonstrates the advantages of integrating in silico analyses with SELEX to identify high-affinity aptamers. The TAGLN2 targeting aptamers identified in this study can potentially be used for the development of diagnostic tests for the detection of TAGLN2. This presents promising opportunities for early detection and intervention in diseases associated with increased levels of TAGLN2 expression.
