ABSTRACT
Salmonella FlgA, a periplasmic protein essential for flagellar P-ring assembly, has been crystallized in two forms. The native protein crystallized in space group C222, with unit-cell parameters a = 107.5, b = 131.8, c = 49.4 Å, and diffracted to about 2.0 Å resolution (crystal form I). In this crystal, the asymmetric unit is likely to contain one molecule, with a solvent content of 66.8%. Selenomethionine-labelled FlgA protein crystallized in space group C222(1), with unit-cell parameters a = 53.2, b = 162.5, c = 103.5 Å, and diffracted to 2.7 Å resolution (crystal form II). In crystal form II, the asymmetric unit contained two molecules with a solvent content of 48.0%. The multiple-wavelength and single-wavelength anomalous dispersion methods allowed the visualization of the electron-density distributions of the form I and II crystals, respectively. The two maps suggested that FlgA is in two different conformations in the two crystals.
Subject(s)
Bacterial Proteins/chemistry , Salmonella/chemistry , Crystallization , Crystallography, X-RaySubject(s)
Bacteria/cytology , Bacterial Physiological Phenomena , Flagella/chemistry , Flagella/physiology , Bacterial Proteins/chemistry , Bacterial Proteins/physiology , Bacterial Proteins/ultrastructure , Crystallography, X-Ray , Flagella/metabolism , Flagella/ultrastructure , Flagellin/chemistry , Flagellin/metabolism , Flagellin/ultrastructure , Microscopy, Electron , Molecular Motor Proteins/chemistry , Molecular Motor Proteins/physiology , Molecular Motor Proteins/ultrastructure , Movement , Protein TransportABSTRACT
A core fragment of the bacterial flagellar hook protein FlgE was overexpressed, purified and crystallized. The crystal diffracted to 1.6 A resolution using synchrotron X-radiation. The crystal belongs to the orthorhombic crystal system, with space group P2(1)2(1)2 and unit-cell parameters a = 128.4, b = 48.8, c = 96.7 A. SeMet protein was also overexpressed, purified, crystallized and a set of 2.3 A MAD data was collected.
Subject(s)
Bacterial Proteins/chemistry , Peptide Fragments/chemistry , Salmonella typhimurium/chemistry , Crystallization , Crystallography, X-RayABSTRACT
The bacterial flagellum is a motile organelle, and the flagellar hook is a short, highly curved tubular structure that connects the flagellar motor to the long filament acting as a helical propeller. The hook is made of about 120 copies of a single protein, FlgE, and its function as a nano-sized universal joint is essential for dynamic and efficient bacterial motility and taxis. It transmits the motor torque to the helical propeller over a wide range of its orientation for swimming and tumbling. Here we report a partial atomic model of the hook obtained by X-ray crystallography of FlgE31, a major proteolytic fragment of FlgE lacking unfolded terminal regions, and by electron cryomicroscopy and three-dimensional helical image reconstruction of the hook. The model reveals the intricate molecular interactions and a plausible switching mechanism for the hook to be flexible in bending but rigid against twisting for its universal joint function.