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Cloning, expression, purification, crystallization and preliminary X-ray diffraction of a lysine-specific permease from Pseudomonas aeruginosa.

DOI: 10.1107/S2053230X14017865 DOI Help
PMID: 25286940 PMID Help

Authors: Emmanuel Nji (Trinity College, Dublin) , Dianfan Li (Trinity College, Dublin) , Declan Doyle (Trinity College, Dublin) , Martin Caffrey (Trinity College, Dublin)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Acta Crystallographica Section F Structural Biology Communications , VOL 70 , PAGES 1362 - 1367

State: Published (Approved)
Published: October 2014
Diamond Proposal Number(s): 7796

Abstract: The prokaryotic lysine-specific permease (LysP) belongs to the amino acid-polyamine-organocation (APC) transporter superfamily. In the cell, members of this family are responsible for the uptake and recycling of nutrients, for the maintenance of a constant internal ion concentration and for cell volume regulation. The detailed mechanism of substrate selectivity and transport of L-lysine by LysP is not understood. A high-resolution crystal structure would enormously facilitate such an understanding. To this end, LysP from Pseudomonas aeruginosa was recombinantly expressed in Escherichia coli and purified to near homogeneity by immobilized metal ion-affinity chromatography (IMAC) and size-exclusion chromatography (SEC). Hexagonal- and rod-shaped crystals were obtained in the presence of L-lysine and the L-lysine analogue L-4-thialysine by vapour diffusion and diffracted to 7.5 Å resolution. The diffraction data were indexed in space group P21, with unit-cell parameters a = 169.53, b = 169.53, c = 290.13 Å, γ = 120°.

Subject Areas: Biology and Bio-materials


Instruments: I24-Microfocus Macromolecular Crystallography

Other Facilities: APS