COMPLEXES OF THE CATALYTIC DOMAIN OF AVIAN SARCOMA VIRUS INTEGRASE WITH DIVALENT CATIONS. Grzegorz Bujacz¹, Mariusz Jaskólski², Jerry Alexandratos¹, Alexander Wlodawer¹, George Merkel³, Richard A. Katz³, Anna Marie Skalka^{3, 1}Macromolecular Structure Laboratory, NCI-Frederick Cancer Research and Development Center, ABL-Basic Research Program, Frederick, MD 21702, ²Center for Biocrystallographic Research, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Pozna_, Poland, ³Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, PA 19111

Retroviral integrases require divalent cations, such as Mg²⁺ or Mn²⁺, for their enzymatic activity. Details of the placement of metal ions in the active site can enhance our understanding of the catalytic mechanism of these enzymes and be helpful in designing integrase inhibitors - potentially potent antiviral drugs. Structures of the enzymatically-active catalytic domain (residues 52-207) of avian sarcoma virus integrase have been solved in the presence of divalent cations Mg²⁺ and Mn²⁺, at resolutions of 1.7 - 2.0 Å. A single ion of either type interacts with the aspartate side chains of the D,D(35)E catalytic center and uses four water molecules to complete its octahedral coordination. Binding of the required metal ions does not lead to significant structural modifications in the active site of ASV IN. Only small adjustments take place in the active site residue side chains upon binding of the metal cofactor. However, the conformation of the catalytic aspartates in the active site of ASV IN (both in the absence or presence of metal) differs significantly from that reported for the analogous residues in HIV-1 IN in the absence of metal. The location of the metal ion is similar to that reported for metal complexes of RNases H from E. coli and HIV-1, which are structurally related to integrase. This suggests that the observed constellation of the acidic residues represents a pre-formed catalytically competent active site. Only a single divalent cation was observed even at extremely high concentrations of the metals. The unexpected differences between the active sites of ASV and HIV-1 IN remain unexplained, but because of ability to create of complex with metal we suggest that at the current stage of investigation, the ASV IN structures provides a good model for inhibitor design.