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

Abstract

This paper presents improved method for isolation of photochrome transmembraine protein bacteriorhodopsin (output 5 mg from 100 g of wet biomass) capable to transform light energy to electrochemical energy of generated protons H⁺ and ??P. The protein was isolated from purple membranes of photo-organotrophic halobacterium Halobacterium halobium 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 elecrtophoresis 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