Co-crystal of Escherichia coli RNase HI with Mn2+ ions reveals two divalent metals bound in the active site

Ribonuclease H (RNase H) selectively degrades the RNA strand of RNA . DNA hybrids in a divalent cation-dependent manner. Previous structural studies revealed a single Mg2+ ion-binding site in Escherichia coli RNase HI. In: the crystal structure of the related RNase H domain: of human immunodeficiency virus reverse transcriptase, however, two Mn2+ ions were observed suggesting a different mode of metal binding. E, coli RNase HI shows catalytic activity in the presence of Mg2+ or Mn2+ ions, but: these two metals show strikingly different optimal concentrations. Mg2+ ions are required in millimolar concentrations, but Mn2+ ions are only required-in micromolar quantities. Based upon the metal dependence off. coli RNase HI activity, we proposed an activation/attenuation model in which one metal is required for catalysis, and binding of a second metal is inhibitory. We have now solved the co-crystal structure of E, coli RNase HI with Mn2+ ions at 1.9-Angstrom resolution. Two octahedrally coordinated Mn2+ ions are seen to bind to the enzyme-active site. Residues Asp-10, Glu-48, and Asp-70 make direct (inner sphere) coordination contacts to the first (activating) metal, whereas residues Asp-10 and Asp-134 make direct contacts to the second (attenuating) metal. This structure is consistent with biochemical evidence suggesting that two metal ions may bind RNase H but liganding a second ion inhibits RNase H activity.