Novel strategy for enhanced expression and purification of recombinant Bacteriorhodopsin in Escherichia coli

Purification of recombinant bacteriorhodopsin (BR) from E.coli was achieved by two step process that includes the expression of Bacterio-opsin (BO, apoprotein) and renaturation with retinol. However, the expression of BO in E.coli was hampered by the extensive cytoplasmic proteolytic degradation. The usage of N-terminal signal sequence particularly, the Mistic sequence was efficient enough to avoid proteolysis and yields a maximum quantity of BO through E.coli expression system. The present investigation was designed to maximize the final yield of BO by optimizing the codons of Mistic and BO sequence according to the codon usage in E. coli expression system. Codon optimized Mistic-BO expression construct was developed and successfully expressed in E.coli, efficiently purified and renatured to obtain a functional BR to a maximum yield of 200 mg/l. The observed yield is highest among the existing reports of recombinant BR produced by overexpression in E.coli system.