Analysis of Halobacterium halobium Gas Vesicles

Abstract

Gas vesicles, isolated from lysed Halobacterium halobium cells, gave an amino acid analysis which accounted for 78% of the weight, and the balance was mainly salt and water. One percent of tightly bound d-galactose was found, as well as 2% of phosphate that was not released by treatment which promotes beta-elimination, by hydrolytic release of the galactose, by carboxymethylation of lysine, or by alkaline phosphatase digestion. Only a trace of lipid was detected, and it appeared to have a polyisoprenoid structure. The vesicles were not solubilized by extremes of pH, by agents such as urea, guanidine hydrochloride, formic acid, and detergents, or by organic solvents. Succinylation and carboxymethylation gave partial dispersion, but the products were heterogeneous and of high molecular weight. The amino acid composition of vesicles was independent of fragment size. No band was obtained by polyacrylamide gel electrophoresis, with neutral, acidic, and alkaline systems, with or without sodium dodecyl sulfate and urea, before or after chemical modification. No amino terminus was detected. Electrofocusing of a vesicle dispersion showed a major component with a pI of 4.0 and an amino acid composition of the whole vesicles, and a minor band with pI 3.4 which had an amino acid composition different from whole vesicles. Vesicle protein was resistant to digestion by Pronase, trypsin, thermolysin, and papain. The precipitin reaction with rabbit antivesicle serum was not inhibited by galactose or inorganic phosphate. Succinylated and carboxymethylated vesicles cross-reacted with antivesicle serum. Cell lysates contained material which reacted with antiserum, but it was heterogeneous and mainly larger than 5 x 10(6) daltons. Material from nonvacuolated mutants reacted weakly with antiserum, but the amino acid composition of the precipitated antigen was different from that of vesicles and of soluble cross-reacting material from vacuolated cells.