Temperate bacteriophage (phage) GIL01 infects insect pathogen Bacillus thuringiensis. In the host cell, the phage can trigger the lytic or the lysogenic cycle. Recent studies are showing that GIL01 genes are transcribed from three different promoters. Lytic promoter P3 is responsible for the synthesis of phage capsid and for the release of phages out of host cell. Promoter P3 is activated by the protein gp6 of which gene is transcribed from phage GIL01 promoter P1 in the DNA damage response. gp6 protein acts as an activator by interacting with the P3 promoter region, covering the -35 promoter element, which enables RNA polymerase to bind to the promoter and activates transcription of late phage genes. With set of primers and with “Phusion” DNA polymerase we introduced point mutations into the gene gp6 carried by the plasmid. We triggered the synthesis of the wild type protein gp6 or its derivates inside the strain B. thuringiensis, which carried also the plasmid with reporter gene lacZ under control of P3promoter. Using the beta-galactosidase assay, we analysed the effect of gp6 protein or its derivates on the activity of P3. We conclude that residues tyrosine 34 (gp6 Y34A) and glutamine 18 (gp6 E18A) are important for the activation of P3 promoter, thus for the activation of the lytic cycle of phage GIL01. With surface plasmon resonance we proved that protein gp6 with point mutations gp6 Y34A and gp6 E18A, bind to the P3 promoter region.