SpaB, an atypically adhesive basal pilin from the lactobacillar SpaCBA pilus: crystallization and X-ray diffraction analysis

Abhin Kumar Megta, Airi Palva, Ingemar von Ossowski, Vengadesan Krishnan*

*Tämän työn vastaava kirjoittaja

Tutkimustuotos: LehtiartikkeliArticleScientificvertaisarvioitu

Abstrakti

The SpaB pilin is recognized as the basal subunit of the sortase-dependent SpaCBA pilus, which is known to be produced by the Gram-positive Lactobacillus rhamnosus GG, a gut-adapted commensal advocated to have health benefits. Despite seeming to function as an archetypal basal pilin by serving as the terminal subunit in pilus assembly, SpaB also assumes an atypical role as a mucoadhesive protein. To shed light on the structural factors that contribute to this dual functional behaviour, a recombinant form of the L. rhamnosus GG SpaB pilin was produced and purified for crystallization and X-ray diffraction experiments. The crystallization of SpaB remained particularly challenging until the implementation of a three-pronged crystallization approach involving C-terminal tail truncation, surface lysine methylation and magnesium additives. Ultimately, hexagonal crystals of SpaB were produced and were able to diffract to a resolution of 2.4 angstrom. This crystal form belonged to space group P6(5)22 or P6(1)22, with unit-cell parameters a = b = 51.53, c = 408.22 angstrom, alpha = beta = 90.0, gamma = 120.0 degrees. Obtaining an interpretable electron-density map via single-wavelength anomalous diffraction (SAD) using iodide-derivative data sets did not succeed owing to the weak anomalous signal. As an alternative, attempts to provide phases by molecular replacement using the iodide-SAD data from SpaB and a collection of distant homology models (

AlkuperäiskieliEnglanti
Sivut731-737
Sivumäärä7
JulkaisuActa crystallographica section F : Structural biology communications
Vuosikerta75
DOI - pysyväislinkit
TilaJulkaistu - jouluk. 2019
OKM-julkaisutyyppiA1 Julkaistu artikkeli, soviteltu

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