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Fusobacterium nucleatum is associated with the incidence and development of multiple diseases, such as periodontitis and colorectal cancer (CRC). Till now, studies have proved only a few proteins to be associated with such pathogenic diseases. The two-component system is one of the most prevalent forms of bacterial signal transduction related to intestinal diseases. Here we report a novel, recombinant, two-component, response-regulator protein ArlR from the genome of F. nucleatum strain ATCC 25586. We optimized the expression and purification conditions of ArlR; in addition, we characterized the interaction of this response regulator protein to the corresponding histidine kinase and DNA sequence. The full-length ArlR was successfully expressed in six of the E. coli host strains. However, optimum expression conditions of ArlR were present only in E. coli strain BL21 Condon plus (DE3) RIL that was later induced with isopropyl β-D-1-thiogalactopyranoside (IPTG) for 8 h at 25 °C. The SDS-PAGE analysis revealed the molecular weight of the recombinant protein as 27.3 kDa and approximately 90% purity after gel filtration chromatography. ArlR was biologically active after its purification. Therefore, it accepted the corresponding phosphorylated histidine-kinase phosphate group and bound to the analogous DNA sequence. The binding constant between ArlR and the corresponding histidine kinase is 1.28 µM, while the binding constant between ArlR and the bound DNA sequence is 37.5 µM. Altogether, these results illustrate an effective expression and purification method for the novel two-component system protein ArlR.

Fusobacterium nucleatum is associated with the incidence and development of multiple diseases, such as periodontitis and colorectal cancer (CRC). Till now, studies have proved only a few proteins to be associated with such pathogenic diseases. The two-component system is one of the most prevalent forms of bacterial signal transduction related to intestinal diseases. Here we report a novel, recombinant, two-component, response-regulator protein ArlR from the genome of F. nucleatum strain ATCC 25586. We optimized the expression and puri cation conditions of ArlR; in addition, we characterized the interaction of this response regulator protein to the corresponding histidine kinase and DNA sequence. The fulllength ArlR was successfully expressed in six of the E. coli host strains. However, optimum expression conditions of ArlR were present only in E. coli strain BL21 Condon plus (DE3) RIL that was later induced with isopropyl β-D-1-thiogalactopyranoside (IPTG) for 8 h at 25 °C. The SDS-PAGE analysis revealed the molecular weight of the recombinant protein as 27.3 kDa and approximately 90% purity after gel ltration chromatography. ArlR was biologically active after its puri cation. Therefore, it accepted the corresponding phosphorylated histidine-kinase phosphate group and bound to the analogous DNA sequence. The binding constant between ArlR and the corresponding histidine kinase is 1.28 µM, while the binding constant between ArlR and the bound DNA sequence is 37.5 µM. Altogether, these results illustrate an effective expression and puri cation method for the novel two-component system protein ArlR.