The flagellar membrane of Ochromonas danica. Isolation and electrophoretic analysis of the flagellar membrane, axonemes, and mastigonemes.

The isolation and purification of the flagellar membrane of the phytoflagellate, Ochromonas danica, is described. The procedure is simple, mild, rapid, and it produces a pure membrane preparation. The method additionally permits the isolation of clean preparations of axonemes and mastigonemes from a single flagella preparation. Each component was studied by electron microscopy and acrylamide gel electrophroesis. The isolated flagella preparation was nearly free of other cellular organelles as judged by phase contrast and electron microscopy. The purified membrane preparation consisted of small vesicles (500 to 1500 A in diameter) with a trilamellar pattern about 80 A thick. Isolated membrane was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, displaying five major protein bands, five minor protein bands, and some protein which remained at the origin. The five major protein components had apparent molecular weights of 54,000, 47,000, 35,000, 31,000, and 28,000. All mastigoneme protein components are glycoproteins as judged by periodic acid-Schiff staining. The mastigoneme preparation contained three major protein bands. Two of them were revealed as doublets and migrated with an average velocity corresponding to 83,000 delatons, the other major protein band migrated with a velocity corresponding to 54,000 daltons. A heavy carbohydrate band is seen near the bromphenol blue tracking dye. The axoneme preparation showed one major protein band having an apparent molecular weight of about 54,000 and some proteins having high molecular weights which remained on top of the polyacrylamide gel.