The Improved Method for Isolation of Photochrome Transmembrane Protein Bacteriorhodopsin from Purple Membranes of Halobacterium Halobacterium Halobium ET 1001

Abstract

We proposed the improved method for isolation of photochrome transmembrane protein bacteriorhodopsin (output 5 mg from 100 g of wet biomass) capable to transform light energy to electrochemical energy of generated protons H⁺ and АТP. The protein was isolated from purple membranes of photo-organotrophic halobacterium Halobacterium halobium ET 1001 by cellular autolysis by distilled water, processing of bacterial biomass by ultrasound at 22 KHz, alcohol extraction of low and high-weight molecular impurities, cellular RNA, carotenoids and lipids, solubilization with 0.5% (w/v) SDS-Na, fractionation by MeOH and column gel permeation chromatography (GPC) of the final protein on Sephadex G-200 with 0.1% (w/v) SDS-Na and 2.5 mM ETDA. The homogeneity of the isolated BR was proved by combination of preparative and analytical methods including electrophoresis in 12.5% (w/v) PAAG with 0.1% (w/v) SDS-Na and regeneration of apomembranes with 13-trans-retinal.

Keywords: Halobacterium halobium, purple membranes, bacteriorhodopsin, biosynthesis, biomolecular electronics