Summary information

PDB id
6eo6
Class
hydrolase-DNA
Method
X-ray (1.69 Å)
Summary
X-ray structure of the complex between human alpha-thrombin and modified 15-mer DNA aptamer containing 5-(3-(2-(1h-indol-3-yl)acetamide-n-yl)-1-propen-1-yl)-2'-deoxyuridine residue
Reference
Dolot R, Lam CH, Sierant M, Zhao Q, Liu FW, Nawrot B, Egli M, Yang X (2018): "Crystal structures of thrombin in complex with chemically modified thrombin DNA aptamers reveal the origins of enhanced affinity." Nucleic Acids Res., 46, 4819-4830. doi: 10.1093/nar/gky268.
Abstract
Thrombin-binding aptamer (TBA) is a DNA 15-mer of sequence 5'-GGT TGG TGT GGT TGG-3' that folds into a G-quadruplex structure linked by two T-T loops located on one side and a T-G-T loop on the other. These loops are critical for post-SELEX modification to improve TBA target affinity. With this goal in mind we synthesized a T analog, 5-(indolyl-3-acetyl-3-amino-1-propenyl)-2'-deoxyuridine (W) to substitute one T or a pair of Ts. Subsequently, the affinity for each analog was determined by biolayer interferometry. An aptamer with W at position 4 exhibited about 3-fold increased binding affinity, and replacing both T4 and T12 with W afforded an almost 10-fold enhancement compared to native TBA. To better understand the role of the substituent's aromatic moiety, an aptamer with 5-(methyl-3-acetyl-3-amino-1-propenyl)-2'-deoxyuridine (K; W without the indole moiety) in place of T4 was also synthesized. This K4 aptamer was found to improve affinity 7-fold relative to native TBA. Crystal structures of aptamers with T4 replaced by either W or K bound to thrombin provide insight into the origins of the increased affinities. Our work demonstrates that facile chemical modification of a simple DNA aptamer can be used to significantly improve its binding affinity for a well-established pharmacological target protein.
G4 notes
2 G-tetrads, 1 G4 helix, 1 G4 stem, 2(+Ln+Lw+Ln), chair(2+2), UDUD

Base-block schematics in six views

PyMOL session file PDB file View in 3Dmol.js

List of 2 G-tetrads

 1 glyco-bond=s-s- sugar=---- groove=wnwn planarity=0.499 type=saddle nts=4 GGGG D.DG401,D.DG415,D.DG410,D.DG406
 2 glyco-bond=-s-s sugar=---- groove=wnwn planarity=0.562 type=other  nts=4 GGGG D.DG402,D.DG414,D.DG411,D.DG405

List of 1 G4-helix

In DSSR, a G4-helix is defined by stacking interactions of G-tetrads, regardless of backbone connectivity, and may contain more than one G4-stem.

Helix#1, 2 G-tetrad layers, INTRA-molecular, with 1 stem

 1  glyco-bond=s-s- sugar=---- groove=wnwn Major-->WC nts=4 GGGG D.DG401,D.DG415,D.DG410,D.DG406
 2  glyco-bond=-s-s sugar=---- groove=wnwn WC-->Major nts=4 GGGG D.DG402,D.DG414,D.DG411,D.DG405
  step#1  mm(<>,outward)  area=16.20 rise=3.51 twist=14.8
  strand#1 DNA glyco-bond=s- sugar=-- nts=2 GG D.DG401,D.DG402
  strand#2 DNA glyco-bond=-s sugar=-- nts=2 GG D.DG415,D.DG414
  strand#3 DNA glyco-bond=s- sugar=-- nts=2 GG D.DG410,D.DG411
  strand#4 DNA glyco-bond=-s sugar=-- nts=2 GG D.DG406,D.DG405

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1 stacking diagram 
 1  glyco-bond=s-s- sugar=---- groove=wnwn Major-->WC nts=4 GGGG D.DG401,D.DG415,D.DG410,D.DG406
 2  glyco-bond=-s-s sugar=---- groove=wnwn WC-->Major nts=4 GGGG D.DG402,D.DG414,D.DG411,D.DG405
  step#1  mm(<>,outward)  area=16.20 rise=3.51 twist=14.8

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List of 1 G4-stem

In DSSR, a G4-stem is defined as a G4-helix with backbone connectivity. Bulges are also allowed along each of the four strands.

Stem#1, 2 G-tetrad layers, 3 loops, INTRA-molecular, UDUD, anti-parallel, 2(+Ln+Lw+Ln), chair(2+2)

 1  glyco-bond=s-s- sugar=---- groove=wnwn Major-->WC nts=4 GGGG D.DG401,D.DG415,D.DG410,D.DG406
 2  glyco-bond=-s-s sugar=---- groove=wnwn WC-->Major nts=4 GGGG D.DG402,D.DG414,D.DG411,D.DG405
  step#1  mm(<>,outward)  area=16.20 rise=3.51 twist=14.8
  strand#1  U DNA glyco-bond=s- sugar=-- nts=2 GG D.DG401,D.DG402
  strand#2  D DNA glyco-bond=-s sugar=-- nts=2 GG D.DG415,D.DG414
  strand#3  U DNA glyco-bond=s- sugar=-- nts=2 GG D.DG410,D.DG411
  strand#4  D DNA glyco-bond=-s sugar=-- nts=2 GG D.DG406,D.DG405
  loop#1 type=lateral   strands=[#1,#4] nts=2 Ta D.DT403,D.77Y404
  loop#2 type=lateral   strands=[#4,#3] nts=3 TGT D.DT407,D.DG408,D.DT409
  loop#3 type=lateral   strands=[#3,#2] nts=2 TT D.DT412,D.DT413

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