Article thumbnail

Direct Detection of Thrombin Binding to 8-Bromodeoxyguanosine-Modified Aptamer: Effects of Modification on Affinity and Kinetics

By Shou Goji and Jun Matsui


The affinity of an 8-bromodeoxyguanosine- (8-BrdG-) substituted thrombin-binding aptamer (TBA-Br), which has the 1st and 10th guanosine residues replaced with 8-BrdG, was estimated using reflectometric interference spectroscopy (RIfS). When comparing TBA-Br with unmodified TBA (TBA-H), it was demonstrated that the modification effectively improved the affinity of TBA; dissociation constants (KD) of TBA-H and TBA-Br were 45.4 nM and 1.99 nM, respectively. These values, which were obtained by direct observation of thrombin binding using RIfS, have the same order of magnitude as those obtained in our previous study utilizing conformational changes in TBA to detect thrombin binding, thus confirming the validity of the obtained KD values. RIfS measurements also revealed that the 8-BrdG modification resulted in a lower dissociation rate constant (kd), which suggests that the enhancement of affinity can be attributed to the stabilization of the G-quadruplex structure on introduction of 8-BrdG

Topics: Research Article
Publisher: SAGE-Hindawi Access to Research
OAI identifier:
Provided by: PubMed Central

To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.

Suggested articles


  1. (2008). a s e g a w a ,K .I .T a i r a ,K .S o d e ,a n dK .I k e b u k u r o ,“ I m -provement of aptamer affinity by dimerization,”
  2. (1996). An ambiguous structure of a DNA 15-mer thrombin complex,”
  3. (2007). and L.Mayol,“A novel thrombin binding aptamer containinga GLNAresidue,”
  4. (1993). ang,S.M cC ur d y ,R.G.Shea,S.S waminathan,andP .H. Bolton, “A DNA aptamer which binds to and inhibits thrombin exhibits a new structural motif for
  5. (1992). B o c k ,L .C .G r i ffin,
  6. (2007). Biophysical properties of quadruplexes containing two or three 8-bromodeoxyguanosine residues,”
  7. (2009). Combinatorialsynthesis ofthrombin-binding aptamers containing iso-guanine,”
  8. (2005). Determination of affinity constants of locked nucleic acid (LNA) and DNA duplex formation using label free sensor technology,”
  9. (2008). Effect of locked-nucleic acid on a biologically active G-quadruplex. A structure-activity relationship of the thrombin aptamer,”
  10. (2008). Electrochemical impedance spectroscopy for study of aptamer-thrombin interfacial interactions,”
  11. (2011). Enhanced anti-HIV-1 activity ofG-quadruplexes comprising locked nucleic acids and intercalating nucleic acids,”
  12. (2007). G-quadruplex induced stabilization by 2 -deoxy-2 -fluoro-d-arabinonucleic acids (2
  13. (2007). h a n g g u a n ,Z .T a n g ,P .M a l l i k a r a t c h y ,Z .X i a o ,a n dW .T a n , “Optimization and modifications of aptamers selected from live cancer cell lines,”
  14. (2010). High affinity anti-inorganic material antibody generation by integrating graft and evolution technologies: potential of antibodies as biointerface molecules,” J o u r n a lo fB i o l o g i c a lC h e m i s t r y ,v o l .
  15. (2007). i a n i k ,V .O s t a t n ´
  16. (1997). i a s ,J .L .B a t t i s t e ,a n dJ .R .W i l l i a m s o n ,“ C h e m i c a lp r o b e forglycosidicconformationin telomericDNAs,”
  17. (2011). Improved thrombin binding aptamer by incorporation of a single unlocked nucleic acid monomer,”
  18. (1997). Label-free monitoring of DNA-ligand interactions,”
  19. (2005). Labelfree characterisation of oligonucleotide hybridisation using reflectometric interference spectroscopy,”
  20. (1993). M a c a y a ,P .S c h u l t z e
  21. (1997). O s b o r n e ,R .J .C a i n ,a n dG .D .G l i c k ,“ S t r u c t u r ea n d dynamics of disulfide cross-linkedDNA triple helices,”
  22. (1998). r a w c z y k ,S .S w a m i n a t h a ne ta l . ,“ N 2-a n dC 8-substituted oligodeoxynucleotides with enhanced thrombin inhibitory activity in vitro and in vivo,”
  23. (2004). S c h m i d t ,S .B o r k o w s k i ,J .K u r r e c ke ta l . ,“ A p p l i c a t i o no f locked nucleic acids to improve aptamer in vivo stability and targeting function,”
  24. (2004). s p o s i t o ,A .R a n d a z z o ,G .P i c c i a l l i ,L .P e t r a c c o n e
  25. (2005). The effect of chemical modifications on the thermal stability of different G-quadruplex-forming oligonucleotides,”
  26. (1993). The structure of α-thrombin inhibited by a 15-mer single-stranded DNA aptamer,”
  27. (1994). Three-dimensional solution structure of the thrombin-binding