Characterization of the self association of avian sarcoma virus integrase by analytical ultracentrifugation

Retroviral integration protein (IN) has been shown to be both necessary and sufficient for the integration of reverse-transcribed retroviral DNA into the host cell DNA. It has been demonstrated that self-assembly of IN is ess ential for proper function. Analytical ultracentrifugation was used to dete rmine the stoichiometry and free energy of self-association of a full-lengt h IN in various solvents at 23.3 degrees C. Below 8% glycerol, an associati on stoichiometry of monomer-dimer-tetramer is observed. At salt concentrati ons above 500 mM, dimer is the dominant species over a wide range of protei n concentrations. However, as physiological salt concentrations are approac hed, tetramer formation is favored. The addition of glycerol to 500 mM NaCl , 20 mM Tris (pH 8.4), 2 mar beta-mercaptoethanol significantly enhances di mer formation with little effect on tetramer formation. Furthermore, as ele ctrostatic shielding is increased by increasing the ionic strength or decre asing the cation size, dimer formation is strengthened while tetramer forma tion is weakened Taken together, the data support a model in which dimer fo rmation includes favorable buried surface interactions which are opposed by charge-charge repulsion, while favorable electrostatic interactions contri bute significantly to tetramer formation.