Manipulation of the dimer interface for photochemical regulation of the activity of an endonuclease BamHI

ORGN 305

Koji Nakayama, Masayuki Endo, and Tetsuro Majima. The Institute of Scientific and Industrial Research, Osaka University, Mihogaoka 8-1, Ibaraki, Osaka, 567-0047, Japan
We describe photochemical control of the enzymatic activity by manipulation of the dimer interface by employing photofunctionalized BamHI mutants. BamHI has H-bondings in the dimer interface and needs dimer formation to show the activity. We introduced unnatural amino acids having photoremovable 2-nitroveratoryl groups into Lys132 in the H-bondings on the dimer interface of the BamHI mutant (H133A). The photofunctionalized BamHI mutant showed no activity and the following photoirradiation induced the recovery of its activity. Removal of the photofunctional group initiated the activity, which corresponds to the recovery of the specific interactions in the dimer interface. Therefore, photofunctional group introduced to the H-bondings induces the misalignment of the two BamHI monomers and removal of photofunctional group rearranges the specific H-bondings in the dimer interface for the correct dimer formation. The key amino acid for control of the activity is the Lys132 which can be available for the potential control of the activity.