Purification, crystallization and preliminary crystallographic analysis of the ligand-binding regions of the PctA and PctB chemoreceptors from Pseudomonas aeruginosa in complex with amino acids

Acta Crystallogr Sect F Struct Biol Cryst Commun. 2013 Dec;69(Pt 12):1431-5. doi: 10.1107/S1744309113023592. Epub 2013 Nov 29.

Abstract

Pseudomonas aeruginosa is an opportunistic pathogen and one of the major model organisms for the study of chemotaxis. The bacterium harbours 26 genes encoding chemoreceptors, most of which have not been annotated with a function. The paralogous chemoreceptors PctA and PctB (Pseudomonas chemotactic transducer A and B) were found to mediate chemotaxis towards L-amino acids. However, the ligand spectrum of the receptors is quite different since the recombinant ligand-binding region (LBR) of PctA binds 17 different L-amino acids whereas that of PctB recognizes only five. To determine the molecular basis underlying this ligand specificity, PctA-LBR and PctB-LBR have been purified and crystals have been produced after pre-incubation with L-Ile and L-Arg, respectively. Initial crystallization conditions have been identified by the counter-diffusion method and X-ray data have been collected at 2.5 Å (PctA-LBR bound to L-Ile) and 3.14 Å (PctB-LBR bound to L-Arg) resolution. Crystals belonged to space groups P2(1)2(1)2(1) and P3(1)2(1), with unit-cell parameters a = 72.2, b = 78.5, c = 116.6 Å and a = b = 111.6, c = 117.4, respectively, for PctA-LBR and PctB-LBR. Molecular-replacement methods will be pursued for structural determination.

Keywords: Pseudomonas aeruginosa; chemoreceptors PctA and PctB.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Arginine / chemistry*
  • Arginine / metabolism
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Chemotaxis / genetics
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Isoleucine / chemistry*
  • Isoleucine / metabolism
  • Ligands
  • Molecular Sequence Data
  • Protein Interaction Domains and Motifs
  • Pseudomonas aeruginosa / chemistry*
  • Pseudomonas aeruginosa / genetics
  • Pseudomonas aeruginosa / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Alignment

Substances

  • Bacterial Proteins
  • Ligands
  • PctA protein, Pseudomonas aeruginosa
  • PctB protein, Pseudomonas aeruginosa
  • Recombinant Proteins
  • Isoleucine
  • Arginine