Retroviral integrase functions as a multimer and can turn over catalytically

Area of Science:

• Biochemistry
• Molecular Biology
• Virology

Background:

• Retroviral integration is a critical step in the viral life cycle.
• The integrase (IN) protein mediates viral DNA integration into the host genome.
• The precise biochemical and biophysical mechanisms of IN activity remain incompletely understood.

Purpose of the Study:

• To investigate the catalytic activity and quaternary structure of Rous sarcoma virus integrase (IN).
• To determine if IN exhibits multiple turnover capabilities in viral DNA processing and joining reactions.
• To elucidate the minimal functional unit of IN required for its enzymatic activities.

Main Methods:

• In vitro enzymatic assays were used to measure IN's processing and joining activities.
• Steady-state kinetic studies were performed to analyze reaction rates.
• Analytical ultracentrifugation was employed to assess IN's oligomeric state.

Main Results:

• Rous sarcoma virus IN demonstrated multiple turnover in both DNA processing and joining reactions.
• The turnover number for the processing reaction was determined to be 0.26 cleavages/min/mol of IN.
• Kinetic and ultracentrifugation data indicated that IN exists in a dynamic equilibrium of monomeric, dimeric, and tetrameric forms, with dimers being the minimal functional unit.

Conclusions:

• Retroviral integrase (IN) is catalytically active in multiple turnover reactions.
• Both processing and joining activities of IN require a multimeric form.
• An IN dimer represents the minimal functional unit for viral DNA end processing and joining during retroviral integration.