Ultrastructural and conformational changes in gas vacuole membranes isolated from Microcystis aeruginosa

Highly purified, intact gas vacuoles were isolated from the blue-green alga Microcystis aeruginosa. The cylindrical gas vacuoles have a constant diameter of 69 nm and a length of 360 ± 90 nm; the vacuolar membrane is 3 nm thick and consists only of protein. Local conformational changes of the vacuolar membrane were investigated with spin- and fluorescence-labeling techniques. Studies on the temperature dependence of the electron paramagnetic resonance (EPR) spectra of spin-labeled intact vacuoles demonstrate a sharply defined transition temperature of 39 °. Below that temperature the conformational change of the vacuolar membrane remains thermally reversible; above that temperature irreversible processes take place. The rate of tumbling of the spin label attached to the vacuolar surface increases concurrently with the temperature, indicating that in the membrane new modes of vibrations are thermally induced in the protein which narrow the line width of EPR spectra. A solution of the intact gas vacuoles, which has a milky appearance, clears upon application of hydrostatic pressure; the EPR spectrum of the spin-labeled membrane becomes more symmetric and the area under the middle hyperfine line is reduced by approximately 25% compared to intact vacuoles. This suggests a rearrangement of the the protein folding of the membrane such that the paramagnetic label is less restricted in its freedom of motion relative to the protein surface. Intact vacuoles labeled with anilinonapththalene sulfonate show a weak fluorescence while vacuoles subjected to pressure fluoresce strongly. Electron micrographs show that vacuoles collapse under pressure and consist of membranous sheets, ribs, and granules. Irreversible conformational changes of the membrane were also induced by guanidine-HCl, chloroform, and trypsin digestion.