Work

Construction of a Z-DNA-specific restriction endonuclease

Year: 1997

Type: article

Abstract: Novel restriction enzymes can be created by fusing the nuclease domain of FokI endonuclease with defined DNA binding domains. Recently, we have characterized a domain (Z alpha) from the N-terminal region of human double-stranded RNA adenosine deaminase (hADAR1), which binds the Z-conformation with high specificity. Here we report creation of a conformation-specific endonuclease, Z alpha nuclease, which is a chimera of Z alpha and FokI nuclease. Purified Z alpha nuclease cleaves negatively supercoiled plasmids only when they contain a Z-DNA forming insert, such as (dC-dG)13. The precise location of the cleavage sites was determined by primer extension. Cutting has been mapped to the edge of the B-Z junction, suggesting that Z alpha nuclease binds within the Z-DNA insert, but cleaves in the nearby B-DNA, by using a mechanism similar to type IIs restriction enzymes. These data show that Z alpha binds Z-DNA in an environment similar to that in a cell. Z alpha nuclease, a structure-specific restriction enzyme, may be a useful tool for further study of the biological role of Z-DNA.

Source: Proceedings of the National Academy of Sciences

DOI: 10.1073/pnas.94.24.12875

Authors: Yang-Gyun Kim, Pearl S. Kim, Alan Herbert, Alexander Rich

Institution: Massachusetts Institute of Technology

Language: English

Topic: DNA and Nucleic Acid Chemistry

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: green

Funders: U.S. Department of Energy, Massachusetts Institute of Technology, National Cancer Institute

Awards: THE MOLECULAR STRUCTURE OF NUCLEIC ACIDS, G8868053602