Structure of DNase I at 2.0 Å resolution suggests a mechanism for binding to and cutting DNA

Bovine pancreatic deoxyribonuclease I (DNase I), an endonuclease that degrades double-stranded DNA in a nonspecific but sequence-dependent manner1–4, has been used as a biochemical tool in various reactions, in particular as a probe for the structure of chromatin and for the helical periodicity of DNA on the nucleosome and in solution5–10. Limited digestion by DNase I, termed DNase I ‘foot-printing’, is routinely used to detect protected regions in DNA–protein complexes11. Recently, we have solved the three-dimensional structure of this glycoprotein (relative molecular mass 30,400) by X-ray structure analysis at 2.5 Å resolution12 and have subsequently refined it crystallographically at 2.0 Å (ref. 26). Based on the refined structure and the binding of Ca2+–thymidine 3′,5′-diphosphate (Ca-pTp) at the active site12, we propose a mechanism of action and present a model for the interaction of DNase I with double-stranded DNA that involves the binding of an exposed loop region in the minor groove of B-DNA and electrostatic interactions of phosphates from both strands with arginine and lysine residues on either side of this loop. We explain DNase I cleavage patterns in terms of this model and discuss the consequences of the extended DNase I–DNA contact region for the interpretation of DNase I footprinting results.