Recognition of a flipped base in a hairpinloop DNA by a small peptide.
Two tiny hairpin DNAs, CORE (dAGGCTTCGGCCT) and AP2 (dAGGCTXCGGCCT; X: abasic nucleotide), fold into almost the same tetraloop hairpin structure with one exception, that is, the sixth thymine (T6) of CORE is exposed to the solvent water (Kawakami, J. et al., Chem. Lett. 2001, 258-259). In the present study, we selected small peptides that bind to CORE or AP2 from a combinatorial pentapeptide library with 2.5 x 10(6) variants. On the basis of the structural information, the selected peptide sequences should indicate the essential qualifications for recognition of the hairpin loop DNA with and without a flipped base. In the selected DNA binding peptides, aromatic amino acids such as histidine for CORE and glutamine/aspartic acid for AP2 were found to be abundant amino acids. This amino acid preference suggests that CORE-binding peptides use pi-pi stacking to recognize the target while hydrogen bonding is dominant for AP2-binding peptides. To investigate the binding properties of the selected peptide to the target, surface plasmon resonance was used. The binding constant of the interaction between CORE and a CORE-binding peptide (HWHHE) was about 1.1 x 10(6) M(-1) at 25 degrees C and the resulting binding free energy change at 25 degrees C (DeltaG degrees (25)) was -8.2 kcal mol(-1). The binding of the peptide to AP2 was also analyzed and the resulting binding constant and DeltaG degrees (25) were about 4.2 x 10(4) M(-1) and -6.3 kcal mol(-1), respectively. The difference in the binding free energy changes (DeltaDeltaG degrees (25)) of 1.9 kcal mol(-1) was comparable to the values reported in other systems and was considered a consequence of the loss of pi-pi stacking. Moreover, the stabilization effect by stacking affected the dissociation step as well as the association step. Our results suggest that the existence of an aromatic ring (T6 base) produces new dominant interactions between peptides and nucleic acids, although hydrogen bonding is the preferable mode of interaction in the absence of the flipping base. These findings regarding CORE and AP2 recognition are expected to give useful information in the design of novel artificial DNA binding peptides.
Kawakami
Junji
J
Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe, Japan. kawakami@konan-u.ac.jp
Okabe
Shinji
S
Tanabe
Yoshiatsu
Y
Sugimoto
Naoki
N
D020650:Combinatorial Chemistry Techniques; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000378:metabolism; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D020349:Surface Plasmon Resonance; D013816:Thermodynamics
D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D019151:Peptide Library; D010455:Peptides; D004247:DNA