Improving soluble expression of β-galactosidase in Escherichia coli by fusion with thioredoxin

Recombinant heterologous proteins can be produced as insoluble aggregates partially or perfectly inactive in Escherichia coli. One of the strateges to improve the solubility of recombinant proteins is fusion with a partner that is excellent in producing soluble fusion proteins. To improve the production of soluble beta-galactosidase, the gene of Thermus thermophilus KNOUC112 beta-galactosidase (KNOUC112 beta-gal) was fused with thioredoxin gene, and optimization of its expression in E. coli TOP10 was performed. KNOUC112 beta-gal in pET-5b was isolated-out, fused with thioredoxin gene in pThioHis C, and transformed to E. coli TOP10. The beta-galactosidase fused with thioredoxin was produced in E. coli TOP10 as dinner and trimer. The productivity of fusion beta-galactosidase expressed via pThioHis C at 37degreesC was about 5 times higher than that of unfused beta-galactosidase expressed via pET-5b at 37degreesC. Inclusion body of beta-galactosidase was formed highly, regardless of the induction by IPTG when KNOUC112 beta-gal was expressed via pET-5b at 37degreesC. Fusion beta-galactosidase expressed at 37degreesC via pThioHis C without the induction by IPTG was soluble, but the induction by IPTG promoted the formation of inclusion body. Lowering the incubation temperature for the expression of fusion gene under 25degreesC prevented the formation of inclusion body, optimally at 25degreesC. 0.07 mM of IPTG was sufficient for the soluble expression of fusion gene at 25degreesC. The soluble production of Thermus thermophilus KNOUC112 beta-galactosidase could be increased about 10 times by fusion with thioredoxin, and optimization of incubation temperature and IPTG concentration for induction.