Кл.слова (ненормированные):
MT: Oligonucleotides: Diagnostics and Biosensors -- COVID-19 diagnosis -- SARS-CoV-2 nucleocapsid detection -- label-free optical aptasensor -- aptamer selection -- biolayer interferometry -- binding motif identification
Аннотация: The spread of COVID-19 has affected billions of people across the globe, and the diagnosis of viral infection still needs improvement. Because of high immunogenicity and abundant expression during viral infection, SARS-CoV-2 nucleocapsid (N) protein could be an important diagnostic marker. This study aimed to develop a label-free optical aptasensor fabricated with a novel single-stranded DNA aptamer to detect the N protein. The N-binding aptamers selected using asymmetric-emulsion PCR-SELEX and their binding affinity and cross-reactivity were characterized by biolayer interferometry. The tNSP3 aptamer (44 nt) was identified to bind the N protein of wild type and Delta and Omicron variants with high affinity (KD in the range of 0.6–3.5 nM). Utilizing tNSP3 to detect the N protein spiked in human saliva evinced the potential of this aptamer with a limit of detection of 4.5 nM. Mass spectrometry analysis was performed along with molecular dynamics simulation to obtain an insight into how tNSP3 binds to the N protein. The identified epitope peptides are localized within the RNA-binding domain and C terminus of the N protein. Hence, we confirmed the performance of this aptamer as an analytical tool for COVID-19 diagnosis.
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Держатели документа:
Department of Chemistry and Biomolecular Sciences, University of Ottawa, Ottawa, ON K1N 6N5, Canada
John L. Holmes Mass Spectrometry Facility, Faculty of Science, University of Ottawa, Ottawa, ON K1N 6N5, Canada
Laboratory for Digital Controlled Drugs and Theranostics, Federal Research Center “Krasnoyarsk Science Center SB RAS”, Krasnoyarsk 660036, Russia
Prof. V.F. Voino-Yasenetsky Krasnoyarsk State Medical University, Krasnoyarsk 660022, Russia
Department of Chemistry, Siberian Federal University, Krasnoyarsk 660041, Russia
Laboratory of Physics of Magnetic Phenomena, Kirensky Institute of Physics, Krasnoyarsk 660036, Russia
Доп.точки доступа:
Poolsup, S.; Zaripov, E.; Huttmann, N.; Minic, Z.; Artyushenko, P. V.; Shchugoreva, I. A.; Tomilin, F. N.; Томилин, Феликс Николаевич; Kichkailo, A. S.; Berezovski, M. V.