Site-specific DNA cleavage by artificial zinc finger-type nuclease with cerium-binding peptide.
The addition of a new function to native proteins is one of the most attractive protein-based designs. In this study, we have converted a C(2)H(2)-type zinc finger as a DNA-binding motif into a novel zinc finger-type nuclease by connecting two distinct zinc finger proteins (Sp1 and GLI) with a functional linker possessing DNA cleavage activity. As a DNA cleavage domain, we chose an analogue of the metal-binding loop (12 amino acid residues), peptide P1, which has been reported to exhibit a strong binding affinity for a lanthanide ion and DNA cleavage ability in the presence of Ce(IV). Our newly designed nucleases, Sp1(P1)GLI and Sp1(P1G)GLI, can strongly bind to a lanthanide ion and show a unique DNA cleavage pattern, in which certain positions between the two DNA-binding sites are specifically cleaved. The present result provides useful information for expanding the design strategy for artificial nucleases.
Nakatsukasa
Takako
T
Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan.
Shiraishi
Yasuhisa
Y
Negi
Shigeru
S
Imanishi
Miki
M
Futaki
Shiroh
S
Sugiura
Yukio
Y
D001483:Base Sequence; D002563:Cerium / Q000378:metabolism*; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D017931:DNA Primers; D003851:Deoxyribonucleases / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D010455:Peptides / Q000378:metabolism*; D016335:Zinc Fingers
D017931:DNA Primers; D010455:Peptides; D002563:Cerium; D004247:DNA; D003851:Deoxyribonucleases