Molecular characterization and prokaryotic expression of buffalo (Bubalus bubalis) Interleukin-6

The study describes the characterization of Interleukin-6 cDNA and essential promoter sequences of the Indian Water Buffalo (Bubalus bubalis) and expression of the recombinant IL-6 in Escherichia coli. Buffalo IL-6 shows very high nucleotide level identity of the cDNA (98.7%) and promoter (98%) sequences with the corresponding cattle sequences. All the major regulatory elements of IL-6 promoter like AP-1, Multiple Response Element, NF-IL6, ETS binding domain and NF-kappaB binding sites show absolute conservation. Basal level IL-6 mRNA is detected in organs like liver, lung and spleen. Concanavalin A stimulated splenocytes produced maximum IL-6 mRNA at 8h poststimulation. Recombinant IL-6 production in JM109 (DE3) and BL21 (DE3) pLysS bacterial system is substantially enhanced by supplementation of rare codon tRNAs through co-transformation with a second plasmid. BL21 (DE3) pLysS strain is a more efficient producer of the IL-6 as it expressed two-fold more protein than by JM109 (DE3) cells. The study shows high-level conservation of IL-6 regulatory and coding sequences between cattle and buffalo, and indicates the use of a common reagent for studying the effects of this cytokine in these species.