AN ESSENTIAL INTERACTION BETWEEN DISTINCT DOMAINS OF HIV-1 INTEGRASE MEDIATES ASSEMBLY OF THE ACTIVE MULTIMER

Integrase mediates integration of the retroviral genome into a host ce ll chromosome, an essential step in the viral life cycle. In vitro, a stable complex containing only purified human immunodeficiency virus ( HIV) integrase and a model viral DNA substrate processively executes t he S'-end processing and DNA joining steps in the integration reaction . We examined the relationship of three essential components of the HI V integrase: the HHCC domain, a putative zinc-finger near the N termin us; the phylogenetically conserved ''DD35E'' motif, which defines the catalytic domain; and a feature recognized by its sensitivity to the a lkylating agent N-ethylmaleimide (NEM). HIV integrase is a multimer, a nd these three components can be distributed among at least two subuni ts of the multimeric enzyme. The components function asymmetrically in the active multimer; the DD35E motif and NEM-sensitive site are requi red in trans to the HHCC region. A divalent cation-dependent interacti on involving the NEM-sensitive site of one integrase subunit and the H HCC region of another subunit points to a role for these two features of integrase in multimer assembly. Deletion of the HHCC domain, or mod ification of integrase with NEM, impaired the assembly of a stable com plex between integrase and viral DNA, suggesting that this initial ste p in the integration pathway requires assembly of the active integrase multimer.