Lin, Y.-W.

ACS Catal. 2020, 10, 14359-14365, 10.1021/acscatal.0c04572

Design of artificial nucleases is essential in biotechnology and biomedicine, whereas few artificial nucleases can both cleave and degrade DNA molecules. Heme proteins are potential enzymes for DNA cleavage. Using a small heme protein, myoglobin (Mb), as a model protein, we engineered a metal-binding motif of [1-His-1-Glu] (native His64 and mutated Glu29) in the heme distal site. The single mutant of L29E Mb was capable of not only efficient DNA cleavage but also DNA degradation upon Mg2+ binding to the heme distal site, as shown by an X-ray crystal structure of the Mg2+-L29E Mb complex. Molecular docking of the protein–DNA complex revealed multiple hydrogen-bonding interactions at their interfaces, involving both minor and major grooves of DNA. Moreover, both the distal Arg45 and the ligand Glu29 were identified as critical residues for the nuclease activity. This study reports the structure of a water-bridged heterodinuclear center of Mg-heme (Mg2+-H2O-Fe3+), showing a similar function as the homodinuclear center (MgA2+-H2O–MgB2+) in natural nuclease, which indicates that the Mg2+-L29E Mb complex is an effective artificial nuclease.