Interaction of the Mnt repressor with 37-base pair synthetic operator DNA fragments. Importance of symmetric GC pairs

Abstract

Mnt repressor is indirectly responsible for the maintenance of lysogeny of the phage P22. This repressor interacts with a 21-base pair operator DNA constituting within it a 17-base pair perfect 2-fold symmetric sequence whose bases make a direct contact with the protein. We have synthesized six 37-base pair DNAs consisting of 21 base pair natural operator and its modifications in which certain symmetrically situated GC base pairs were replaced systematically with ATs to understand their importance. The binding interaction studies of Mnt repressor to such natural and modified operator DNAs reported here indicate that the GCs close to the center of symmetry make major contacts with the protein whereas, GCs nearer to the periphery form weak contacts. Methylation protection experiments indicated that when the GCs near the center of symmetry were replaced with AT, the central GC became more accessible for dimethyl sulfate methylation with possible conformational change in DNA. The circular dichroism studies indicated that upon repressor binding conformational changes in DNA takes place with a possible increase in helicity of the repressor protein.