Bacteria swim by means of a specialized organelle, the bacterial flagellum, which consists of a rotary motor and a propellor. The propellor is a helically shaped filament made up of flagellin subunits. Rotation of the motor turns the filament which in turn propels the cell. Reversal of the motor causes the pitch and hand of the helical filament to change. This structural change, which produces a twiddling or tumbling of the cell, is the mechanism by which the cell changes direction. The combine techniques of electron microscopy and image analysis will be used to study the changes that take place in quaternary structure when the filament changes hand. A three-dimensional image of the filament from Salmonella typhimurium in the two forms will be calculated. These images will provide insights into the mechanics of the structural change and into the structure and packing of the flagellin subunit in the filament. The identity of the flagellar motor is not known, although many consider the basal body an important element. Consisting of a central rod and 4 or 5 rings, the basal body passes from the outer to inner membrane of the cell and serves to anchor the filament to the cell. It is attached to the filament by means of a flexible coupler, the hook. The basal body from Caulobacter crescentus consists of six different proteins. An important goal of this proposal is the identification of these proteins in terms of which genes encode for them and which morphological features they give rise to. Immunoelectron microscopy of basal bodies decorated by antibodies made against individual basal body proteins will be used to locate the proteins. The same antibodies will be used to determine the corresponding gene. Electron microscopy and image analysis will also be used in combination with gel electrophoresis in order to determine the number of subunits of each component in the basal body and to produce a three-dimensional map of the cylindrically averaged basal body structure. A ring of studs surrounding the basal body is found in the inner membrane of the Aquaspirillum serpens cell. It has been suggested that this structure is an important element of the motor. Freeze-fracture, deep-etch, shadowed replicas of Aquaspirillum serpens and Salmonella typhimurium will be used to visualize the structure.
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