Creation and characteristics of unnatural CysHis(3)-type zinc finger protein.
To investigate the properties of unnatural zinc finger peptides with CysHis(3)-type ligand combinations, the HCHH- and CHHH-type zinc finger proteins (zf(HCHH) and zf(CHHH), respectively) were created by mutating Cys to His in the Cys(2)His(2)-type zinc finger of the transcription factor Sp1 (zf(CCHH)). The CD measurements clearly show that the single-finger CysHis(3)-type zinc finger peptides (zf(HCHH)f2 and zf(CHHH)f2) are folded by complexation with Zn(II). From the gel mobility shift assays, the CysHis(3)-type zinc finger proteins (zf(HCHH) and zf(CHHH)) evidently bind to the GC-box DNA, though the DNA binding affinity is lower than that of the wild CCHH-type zinc finger protein. Furthermore, the zf(HCHH)f2 and zf(CHHH)f2 peptides catalyze the hydrolysis of the 4-nitrophenyl acetate in contrast to the catalytically inactive zf(CCHH) peptide. This is the first study of the CysHis(3)-type zinc finger proteins and also provides useful information for redesigning artificial metalloproteins.
Negi
Shigeru
S
Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan.
Itazu
Masako
M
Imanishi
Miki
M
Nomura
Akiko
A
Sugiura
Yukio
Y
D000595:Amino Acid Sequence; D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D006639:Histidine / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D009596:Nitrophenols / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D016329:Sp1 Transcription Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet; D016335:Zinc Fingers
D009596:Nitrophenols; D010446:Peptide Fragments; D016329:Sp1 Transcription Factor; D006639:Histidine; C008642:4-nitrophenyl acetate; D003545:Cysteine