RADIAL MASS ANALYSIS OF THE FLAGELLAR FILAMENT OF SALMONELLA - IMPLICATIONS FOR THE SUBUNIT FOLDING

X-ray fiber diffraction patterns of the R-type straight flagellar fila ment of Salmonella typhimurium SJW1655 strain showed layer-lines with an axial spacing of 1/437 Angstrom(-1), which could be resolved only d ue to very small disorientation angles (<2 degrees) of the filaments i n oriented sol specimens. Although the equatorial layer-line was situa ted between the relatively strong first layer-lines right above and be low it, these small disorientation angles and a new method of two-dime nsional angular deconvolution allowed us to determine the equatorial l ayer-line intensities quite accurately. The equatorial data were phase d by using the amplitude difference between the native flagellar filam ent and its heavy atom derivatives. One of the heavy-atom derivatives was prepared by introducing a cysteine residue by site-directed mutage nesis and applying a mercury compound. From the equatorial structure f actors, the radial density distribution of the filament was calculated at 11 Angstrom resolution. A prominent feature was two pairs of high density peaks at radii of around 25 and 45 Angstrom and a deep density trough between them, which corresponds to the concentric double tubul ar structure in the core region that has been found in the density map recently deduced by helical image reconstruction from electron microg raphs of frozen hydrated filaments. The molecular masses were estimate d for four radial segments that correspond to the morphological domain s identified in the map of helical image reconstruction. Then the doma ins were assigned to sequence positions by correlating the estimated m asses with those of proteolytic fragments of flagellin. The assignment is consistent with the distributions of secondary structures and in p articular a-helical coiled-coils that were predicted from the sequence . It also helps to understand how the polymerization behaviour is affe cted by truncation of the disordered terminal regions of flagellin and why mutations in a specific region are responsible for changes in the polymorphic shape of the filament. (C) 1995 Academic Press Limited