Aptamers are short,single-stranded DNA or RNA molecules that selectively bind to a target molecule.Aptamercomplement duplex(ACD)is often used to design molecular switches capable of producing a detectable signal or tr...Aptamers are short,single-stranded DNA or RNA molecules that selectively bind to a target molecule.Aptamercomplement duplex(ACD)is often used to design molecular switches capable of producing a detectable signal or triggering a structural change upon aptamer binding to a target.However,such aptamer switch generally faces an increased dissociation constant(Kd)due to the energy barrier of the complementary duplex.We reported a competitive hybridization mechanism to modulate the binding affinity of an ACD to a target adenosine.Using the computation-guided design,we calculated the aptamer folding energy for the duplex length from 11-nt to 15-nt,and experimentally measured increased apparent Kd values resulted from these extended duplexes.Using a set of strands to compete with the ACD hybridization,it reduced the aptamer folding energy to facilitate aptamer switches with decreased apparent Kd values ranging from over 400μM without a competing strand to~30μM with a competing strand.This competitive aptamer switch was also found sensitive to single-nucleotide mutations of a competing strand.Our work provides an approach to modulate the binding affinity and the sensitivity of aptamer-complement duplexes,which could be useful in the nucleic acids-based sensing and nanomedicine.展开更多
基金supported by a PECASE award to J.F.(W911NF1910240),a DoD DURIP(W911NF2010107)and a NSF MRI(2215917)J.F.is also supported by the Rutgers TechAdvance award,NJACTS-Pilot Program and NSF PFI award(2141141)D.P.and S.L.appreciate the support of Arts and Sciences Undergraduate Research Grant from the College of Arts and Sciences at Rutgers University-Camden.The authors are grateful to the Equipment Leasing Funds from the State of New Jersey.
文摘Aptamers are short,single-stranded DNA or RNA molecules that selectively bind to a target molecule.Aptamercomplement duplex(ACD)is often used to design molecular switches capable of producing a detectable signal or triggering a structural change upon aptamer binding to a target.However,such aptamer switch generally faces an increased dissociation constant(Kd)due to the energy barrier of the complementary duplex.We reported a competitive hybridization mechanism to modulate the binding affinity of an ACD to a target adenosine.Using the computation-guided design,we calculated the aptamer folding energy for the duplex length from 11-nt to 15-nt,and experimentally measured increased apparent Kd values resulted from these extended duplexes.Using a set of strands to compete with the ACD hybridization,it reduced the aptamer folding energy to facilitate aptamer switches with decreased apparent Kd values ranging from over 400μM without a competing strand to~30μM with a competing strand.This competitive aptamer switch was also found sensitive to single-nucleotide mutations of a competing strand.Our work provides an approach to modulate the binding affinity and the sensitivity of aptamer-complement duplexes,which could be useful in the nucleic acids-based sensing and nanomedicine.
