Highly efficient endonucleolytic cleavage of RNA by a Cys2His2 zinc-finger peptide

We have identified a 30-aa peptide that efficiently cleaves single-stranded RNA. The peptide sequence corresponds to a single zinc finger of the human male-associated ZFY protein; a transcription factor belonging to the Cys(2)His(2) family of zinc-finger proteins, RNA cleavage was observed only in the absence of zinc, Coordination with zinc resulted in complete loss of ribonuclease activity. The ribonuclease active structure was determined to be a homodimeric form of the peptide. Dimerization of the peptide occurred through a single intermolecular disulfide between two of the four cystines, The observed hydrolytic activity was single-stranded RNA-specific, Single-stranded DNA, doublestranded RNA and DNA, and 2'-methoxy-modified sequences were not degraded by the peptide. The peptide specifically cleaved pyrimidines within single-stranded RNA and the dinucleotide sequence 5'-pyr-A-3' was preferred, The RNA cleavage products consisted of a 3' phosphate and 5' hydroxyl, The initial rates of cleavage (V-0) observed for the finger peptide were comparable to rates observed for human ribonucleases, and the catalytic rate (K-cat) was comparable to rates observed for the group II intron rybozymes. The pH profile exhibited by the peptide is characteristic of general acid-base catalytic mechanisms observed with other ribonucleases. These observations raise interesting questions about the potential biological roles of zinc-finger proteins.