Assembly characteristics of flagellar cap protein HAP2 of Salmonella: decamer and pentamer in the pH-sensitive equilibrium.
The cap of the bacterial flagellum is an oligomeric assembly of HAP2 protein (also called FliD), tightly attached to the tip of the flagellar filament. Flagellar growth does not occur in fliD-deficient mutants because flagellin monomers transported through the central channel of the flagellum leak out without polymerizing at the distal end. The structure of the cap complex is not known yet. An in vitro assembly of HAP2 proteins was found to have a pentagonal shape, while its molecular mass corresponded roughly to that of a dodecamer. To characterize the structure and assembly behavior of the complex formed in vitro in more detail, the stoichiometry of the complex and the association equilibrium have been studied. Crosslinking experiments now clearly show that the HAP2 complex is decameric. The assembly equilibrium is mainly between the monomer and decamer with a minor population of intermediate oligomers involved, and is highly dependent on the solution pH as well as the salt concentration: the fraction of the decamer sharply rises as the pH decreases from 8.5 to 8.0; the physiological concentration of salt partially suppresses the decamer formation. A preferential crosslinking within a pentameric unit together with a bipolar feature of the complex particle observed by electron microscopy suggests that the decamer is a bipolar pair of pentamers. Because of the polar nature of the filament cap structure, the pentamer is suggested to be the cap complex with its decamer forming surface involved in interactions with the filament.
Imada
K
K
International Institute for Advanced Research, Matsushita Electric Industrial Co., Ltd., 3-4 Hikaridai, Seika 619-02, Japan.
Vonderviszt
F
F
Furukawa
Y
Y
Oosawa
K
K
Namba
K
K
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents; D004926:Escherichia coli / Q000235:genetics; D005407:Flagella / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012475:Salmonella / Q000235:genetics / Q000378:metabolism*; D012996:Solutions
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D012996:Solutions; C035832:hook protein, bacterial flagellum; C076733:FlaD protein, Bacteria