Isolation and functional expression of the bop gene from Halobiforma lacisalsi

Abstract

A novel bop gene was described from Halobiforma lacisalsi strain AJ5(T), an extremely halophilic archaeon isolated from Ayakekum Lake, China. Following six rounds of PCR amplification based on the conserved fragment of the bop gene, the complete sequence of the bop gene, including the 5' and 3' flanking regions of the conserved fragment, was obtained by the ligation-mediated PCR amplification (LPA) approach. The data presented provide us with further insight into the distribution of bop-like genes in the family Halobacteriaceae. This is the first example of a bop-like gene in halophiles found in the high-pH environment. Alignment and hydropathy analysis of the deduced amino acid sequence identified the conserved functional sites as well as some variations compared with other bacterio-opsins. Molecular phylogenetic analysis revealed the position of the bacterio-opsin of strain AJ5, which is closest to that of Haloterrigena sp. arg-4 with 85% identity. In the presence of all-trans retinal, recombinant Escherichia coli cells expressing the gene turned dark purple. The purple membrane from the recombinant E. coli showed maximal absorption at 540 nm.