Reconstitution of bacterial flagella in vitro

Abstract

The process by which bacterial flagellar filaments are reconstituted from flagellin molecules in vitro is examined by physical measurements (flow birefringence, viscosity, sedimentation) and by electron microscopic analysis, using flagella isolated from a strain of Salmonella. It is shown to be essentially reversible and characteristic of crystallization. In the presence or absence of salt, the flagella are depolymerized into monomers by heat treatment. At neutral pH, the repoly-merization takes place only in the presence of salts. In monomer solutions, however, spontaneous nucleation rarely happens and the solutions remain in a state of super-saturation. In order to polymerize the monomers in such solutions, it is necessary to add fragmented flagella. Then, the ends of added fragments act as nuclei, resulting in rapid formation of long flagellar filaments. In this process the number of filaments contained in each solution remains unchanged, and a one-to-one correspondence holds between the added fragments and the fully grown filaments. The rate of growth of flagellar filaments in vitro is determined under various salt conditions and found to be comparable to that observed in vivo.