Abstract
Aptamers are short single-stranded DNA or RNA oligonucleotides capable of binding with high affinity and specificity to target molecules. Because of their durability and ease of synthesis, aptamers are used in a wide range of biomedical fields, including the diagnosis of diseases and targeted delivery of therapeutic agents. The aptamers were selected using a process called systematic evolution of ligands by exponential enrichment (SELEX), which has been improved for various research purposes since its development in 1990. In this protocol, we describe a modified SELEX method that rapidly produces high aptamer screening yields using two types of magnetic beads. Using this method, we isolated an aptamer that specifically binds to an antimicrobial peptide. We suggest that by conjugating a small therapeutic-specific aptamer to a gold nanoparticle-based delivery system, which enhances the stability and intracellular delivery of peptides, aptamers selected by our method can be used for the development of therapeutic agents utilizing small therapeutic peptides.
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References
Aquino-Jarquin, G. and Toscano-Garibay, J.D. 2011. RNA aptamer evolution: two decades of SELEction. Int. J. Mol. Sci. 12, 9155–9171.
Blind, M. and Blank, M. 2015. Aptamer selection technology and recent advances. Mol. Ther. Nucleic Acids 4, e223.
Camacho, C., Coulouris, G., Avagyan, V., Ma, N., Papadopoulos, J., Bealer, K., and Madden, T.L. 2009. BLAST+: architecture and applications. BMC Bioinformatics 10, 421.
Cibiel, A., Dupont, D.M., and Ducongé, F. 2011. Methods to identify aptamers against cell surface biomarkers. Pharmaceuticals 4, 1216–1235.
Darmostuk, M., Rimpelova, S., Gbelcova, H., and Ruml, T. 2015. Current approaches in SELEX: an update to aptamer selection technology. Biotechnol. Adv. 33, 1141–1161.
Deslouches, B., Gonzalez, I.A., DeAlmeida, D., Islam, K., Steele, C., Montelaro, R.C., and Mietzner, T.A. 2007. De novo-derived cationic antimicrobial peptide activity in a murine model of Pseudomonas aeruginosa bacteraemia. J. Antimicrob. Chemother. 60, 669–672.
Ellington, A.D. and Szostak, J.W. 1990. In vitro selection of RNA molecules that bind specific ligands. Nature 346, 818–822.
Ellington, A.D. and Szostak, J.W. 1992. Selection in vitro of single-stranded DNA molecules that fold into specific ligand-binding structures. Nature 355, 850–852.
Huang, Y.F., Shangguan, D., Liu, H., Phillips, J.A., Zhang, X., Chen, Y., and Tan, W. 2009. Molecular assembly of an aptamer-drug conjugate for targeted drug delivery to tumor cells. Chembiochem 10, 862–868.
Jenison, R.D., Gill, S.C., Pardi, A., and Polisky, B. 1994. High-resolution molecular discrimination by RNA. Science 263, 1425–1429.
Jing, M. and Bowser, M.T. 2011. Isolation of DNA aptamers using micro free flow electrophoresis. Lab Chip 11, 3703–3709.
Kong, H.Y. and Byun, J. 2013. Nucleic acid aptamers: new methods for selection, stabilization, and application in biomedical science. Biomol. Ther. 21, 423–434.
Lee, B., Park, J., Ryu, M., Kim, S., Joo, M., Yeom, J.H., Kim, S., Park, Y., Lee, K., and Bae, J. 2017. Antimicrobial peptide-loaded gold nanoparticle-DNA aptamer conjugates as highly effective antibacterial therapeutics against Vibrio vulnificus. Sci. Rep. 7, 13572.
Luo, Y.L., Shiao, Y.S., and Huang, Y.F. 2011. Release of photoactivatable drugs from plasmonic nanoparticles for targeted cancer therapy. ACS Nano 5, 7796–7804.
Mahlapuu, M., Håkansson, J., Ringstad, L., and Björn, C. 2016. Antimicrobial peptides: an emerging category of therapeutic agents. Front. Cell. Infect. Microbiol. 6, 194.
Makabenta, J.M.V., Nabawy, A., Li, C.H., Schmidt-Malan, S., Patel, R., and Rotello, V.M. 2021. Nanomaterial-based therapeutics for antibiotic-resistant bacterial infections. Nat. Rev. Microbiol. 19, 23–36.
Mendonsa, S.D. and Bowser, M.T. 2004. In vitro selection of high-affinity DNA ligands for human IgE using capillary electrophoresis. Anal. Chem. 76, 5387–5392.
Nimjee, S.M., Rusconi, C.P., and Sullenger, B.A. 2005. Aptamers: an emerging class of therapeutics. Annu. Rev. Med. 56, 555–583.
Park, J., Shin, E., Yeom, J.H., Choi, Y., Joo, M., Lee, M., Kim, J.H., Bae, J., and Lee, K. 2022. Gold nanoparticle-DNA aptamer-assisted delivery of antimicrobial peptide effectively inhibits Acinetobacter baumannii infection in mice. J. Microbiol. 60, 128–136.
Rognes, T., Flouri, T., Nichols, B., Quince, C., and Mahé, F. 2016. VSEARCH: a versatile open source tool for metagenomics. PeerJ 4, e2584.
Ryu, M., Park, J., Yeom, J.H., Joo, M., and Lee, K. 2021. Rediscovery of antimicrobial peptides as therapeutic agents. J. Microbiol. 59, 113–123.
Setlem, K., Mondal, B., Ramlal, S., and Kingston, J. 2016. Immuno affinity SELEX for simple, rapid, and cost-effective aptamer enrichment and identification against aflatoxin B1. Front. Microbiol. 7, 1909.
Song, Y., Zhu, Z., An, Y., Zhang, W., Zhang, H., Liu, D., Yu, C., Duan, W., and Yang, C.J. 2013. Selection of DNA aptamers against epithelial cell adhesion molecule for cancer cell imaging and circulating tumor cell capture. Anal. Chem. 85, 4141–4149.
Stoltenburg, R., Reinemann, C., and Strehlitz, B. 2005. FluMag-SELEX as an advantageous method for DNA aptamer selection. Anal. Bioanal. Chem. 383, 83–91.
Tuerk, C. and Gold, L. 1990. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science 249, 505–510.
Wan, Y., Kim, Y., Li, N., Cho, S.K., Bachoo, R., Ellington, A.D., and Iqbal, S.M. 2010. Surface-immobilized aptamers for cancer cell isolation and microscopic cytology. Cancer Res. 70, 9371–9380.
Yang, Y., Yang, D., Schluesener, H.J., and Zhang, Z. 2007. Advances in SELEX and application of aptamers in the central nervous system. Biomol. Eng. 24, 583–592.
Yazdian-Robati, R., Ramezani, M., Khedri, M., Ansari, N., Abnous, K., and Taghdisi, S.M. 2017. An aptamer for recognizing the transmembrane protein PDL-1 (programmed death-ligand 1), and its application to fluorometric single cell detection of human ovarian carcinoma cells. Microchim. Acta 184, 4029–4035.
Yeom, J.H., Lee, B., Kim, D., Lee, J., Kim, S., Bae, J., Park, Y., and Lee, K. 2016. Gold nanoparticle-DNA aptamer conjugate-assisted delivery of antimicrobial peptide effectively eliminates intra-cellular Salmonella enterica serovar Typhimurium. Biomaterials 104, 43–51.
Zhang, J., Kobert, K., Flouri, T., and Stamatakis, A. 2014. PEAR: a fast and accurate Illumina paired-end reAd mergeR. Bioinformatics 30, 614–620.
Zhou, J. and Rossi, J. 2017. Aptamers as targeted therapeutics: current potential and challenges. Nat. Rev. Drug Discov. 16, 181–202.
Zhuo, Z., Yu, Y., Wang, M., Li, J., Zhang, Z., Liu, J., Wu, X., Lu, A., Zhang, G., and Zhang, B. 2017. Recent advances in SELEX technology and aptamer applications in biomedicine. Int. J. Mol. Sci. 18, 2142.
Acknowledgements
This work was supported by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture and Forestry (IPET) through Animal Disease Management Technology Advancement Support Program, funded by Ministry of Agriculture, Food and Rural Affairs (MAFRA) [grant number: 122060-02-1-CG000 to J. B.] and the National Research Foundation of Korea [NRF-2021R1A2C3008934 to K. L]. This research was also supported by Chung-Ang University Graduate Research Scholarship grants in 2020.
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Lee, J., Ryu, M., Bae, D. et al. Development of DNA aptamers specific for small therapeutic peptides using a modified SELEX method. J Microbiol. 60, 659–667 (2022). https://doi.org/10.1007/s12275-022-2235-4
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DOI: https://doi.org/10.1007/s12275-022-2235-4