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Expression, crystallization and preliminary X-ray diffraction studies on the complete choline-binding domain of the major pneumococcal autolysin.

Carlos Fernández-Tornero·Alvaro Ramón·Concha Fernández-Cabrera·Guillermo Giménez-Gallego·Antonio Romero
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Abstract

The major pneumococcal autolysin (LytA), a virulence factor of this bacterium, is composed of an amino-terminal catalytic domain plus a carboxyl-terminal choline-binding domain (ChBD). This C-terminal domain, responsible for anchorage to the cell wall, is a tandem of six imperfect 20-residue repeats whose precise ends have been difficult to establish by sequence methods. The reported crystal structure of a shortened C-terminal fragment of the protein suggested that it might contain an additional repeat and thus an additional choline-binding site (ChBS). The complete recombinant choline-binding domain of LytA has now been overexpressed in soluble form using a secreting Escherichia coli strain which facilitates purification with a higher yield. It has been crystallized at room temperature using MPD as the main precipitant. The crystals belong to space group P2(1) and diffract to beyond 3.2 A resolution on a synchrotron-radiation source. The molecular-replacement solution indicates that a new ChBS which fits the topology of the solenoid structure is formed in the N-terminal region.

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Binding SitesCholineCrystallizationCrystallography, X-RayModels, MolecularN-Acetylmuramoyl-L-alanine AmidaseProtein ConformationProtein Structure, TertiaryRecombinant ProteinsStreptococcus pneumoniaeJournal ArticleResearch Support, Non-U.S. Gov't

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Carlos Fernández-Tornero, Alvaro Ramón, Concha Fernández-Cabrera, Guillermo Giménez-Gallego, Antonio Romero. 2002-02-21. Expression, crystallization and preliminary X-ray diffraction studies on the complete choline-binding domain of the major pneumococcal autolysin.. https://doi.org/10.1107/s0907444902000537

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