Isoelectric forms of bacteriorhodopsin from Halobacterium halobium

Isoelectric focusing was performed on bacteriorhodopsin isolated from Halobacterium halobium, R₁ strain, solubilized in Nonidet P-40, on sucrose density gradient stabilized columns. When 560 nm absorbance was monitored, four forms of bacteriorhodopsin were observed, having isoelectric points (pI) of: A, 3.93; B, 4.34; C, 5.03; D, 5.49. Instability of some of the isoelectic forms during the very process of electrofocusing was observed. When focused over a period of 6 days, the relative abundance of the forms changed although their pI values remained constant. Refocusing of the isolated forms A or B led to the production of form C. The latter species was stable to refocusing. Form B was unstable to storage either as an aqueous suspension of the purple membrane or as a detergent extract. Each of the forms had the absorption spectrum typical for bacteriorhodopsin and each showed identical patterns after sodium dodecyl sulfate-polyacrylamide gel electrophoresis.