The wild-type repressor CI of temperate mycobacteriophage L1 and the temperature-sensitive (ts) repressor CIts391 of a mutant L1 phage, L1cIts391, have been separately overexpressed in E. coli. Both these repressors were observed to specifically bind with the same cognate operator DNA. The operator-binding activity of CIts391 was shown to differ significantly than that of the CI at 32 to $42^{circ}C$. While 40-95% operator-binding activity was shown to be retained at 35 to $42^{circ}C$ in CI, more than 75% operator-binding activity was lost in CIts391 at 35 to $38^{circ}C$, although the latter showed only 10% less binding compared to that of the former at $32^{circ}C$. The CIts391 showed almost no binding at $42^{circ}C$. An in vivo study showed that the CI repressor inhibited the growth of a clear plaque former mutant of the L1 phage more strongly than that of the CIts391 repressor at both 32 and $42^{circ}C$. The half-life of the CIts391-operator complex was found to be about 8 times less than that of the CI-operator complex at $32^{circ}C$. Interestingly, the repressor-operator complexes preformed at $0^{circ}C$ have shown varying degrees of resistance to dissociation at the temperatures which inhibit the formation of these complexes are inhibited. The CI repressor, but not that of CIts391, regains most of the DNA-binding activity on cooling to $32^{circ}C$ after preincubation at 42 to $52^{circ}C$. All these data suggest that the 131st proline residue at the C-terminal half of CI, which changed to leucine in the CIts391, plays a crucial role in binding the L1 repressor to the cognate operator DNA, although the helix-turn-helix DNA-binding motif of the L1 repressor is located at its N-terminal end.