Department of Biochemistry, Bose Institute, P 1/12, C.I.T. Scheme VIIM, Kolkata-700054, India.
Repressor proteins of temperate mycobacteriophages are distinct from those of lambda and related phages. They bind to asymmetric operator DNAs (located at multiple places in phage genomes) and regulate not only transcription initiation but also transcription elongation. To study the structure-function relationship of mycobacteriophage-specific repressors, we undertook the repressor of temperate mycobacteriophage L1 as a model system and purified it to near homogeneity. We have demonstrated that L1 repressor possesses two domains (namely, N-terminal and C-terminal domains) which are separated by a small hinge region. C-terminal domain is relatively more stable than N-terminal domain. Both intact and C-terminal domain contain significant amounts of a-helix, ß sheet and coils. While intact repressor exists as a mixture of both dimer and monomer in solution, C-terminal domain exists mainly as dimer under identical condition. Intact repressor binds to different L1 operator DNAs with variable affinities and binding is temperature sensitive. Several mutant L1 repressors carrying P131L, E36K, E39Q mutations, respectively, were generated and purified to homogeneity. Among the above mutant repressors, structure and function of P131L significantly differ from those of wild-type repressor. In addition we have also identified several residues of operator DNA which interact with both wild-type and mutant repressors.
