Temperature-dependent Expression of Flagella of Listeria manocytogenes Studied by Electron Microscopy, SDS-PAGE and Western Blotting

Abstract

SUMMARY: Washed cells of Listeria monocytogenes serotype 4b, grown in broth culture at 20°C and at 37°C, were examined by electron microscopy for the presence of flagella. Many flagella were seen in cells grown at 20°C, whereas at 37°C very few were expressed. Flagella sheared from the cell surface were partially purified by differential centrifugation. Using SDS-PAGE and Western blotting two distinct protein bands were seen in this preparation, both with an apparent molecular mass of approximately 29 kDa. Further purification of these proteins was achieved by gel filtration and ion-exchange chromatography. Whole organisms grown at 20°C and 37°C were examined in Western blots using an affinity-purified polyclonal antibody, and a monoclonal antibody, both directed against 29 kDa putative flagellin. Bacteria grown at 20°C expressed abundant flagellin, whereas only trace amounts could be detected in organisms grown at 37°C. It is concluded that organisms grown at 20°C both produce and assemble flagellin at the cell surface, and that flagellin production is a less marked feature of organisms grown at 37°C.