Solution structure and properties of AlgH fromPseudomonas aeruginosa

Abstract

In Pseudomonas aeruginosa, the algH gene regulates the cellular concentrations of a number of enzymes and the production of several virulence factors, and is suggested to serve a global regulatory function. The precise mechanism by which the algH gene product, the AlgH protein, functions is unknown. The same is true for AlgH family members from other bacteria. In order to lay the groundwork for understanding the physical underpinnings of AlgH function, we examined the structure and physical properties of AlgH in solution. Under reducing conditions, results of NMR, electrophoretic mobility, and sedimentation equilibrium experiments indicate AlgH is predominantly monomeric and monodisperse in solution. Under non-reducing conditions intra- and intermolecular disulfide bonds form, the latter promoting AlgH oligomerization. The high-resolution solution structure of AlgH reveals alpha/beta-sandwich architecture fashioned from ten beta strands and seven alpha helices. Comparison with available structures of orthologues indicates conservation of overall structural topology. The region of the protein most strongly conserved structurally also shows the highest amino acid sequence conservation and, as revealed by hydrogen-deuterium exchange studies, is also the most stable. In this region, evolutionary trace analysis identifies two clusters of amino acid residues with the highest evolutionary importance relative to all other AlgH residues. These frame a partially solvent exposed shallow hydrophobic cleft, perhaps identifying a site for intermolecular interactions. The results establish a physical foundation for understanding the structure and function of AlgH and AlgH family proteins and should be of general importance for further investigations of these and related proteins.

CowN sustains nitrogenase turnover in the presence of the inhibitor carbon monoxide

“…The functional importance of CooA regulation of algH is unclear. AlgH is a regulatory protein with an α/β structure of unknown function ( 34 ) and algH is not found in the genomic vicinity of CowN in many diazotrophs. CowN expression is also dependent on the central nitrogenase regulator NifA, which binds to a TGT-(N) 10 -ACA UAS ( 35 ) 59 bp upstream of CowN.…”

Response of Pseudomonas aeruginosa to the Innate Immune System-Derived Oxidants Hypochlorous Acid and Hypothiocyanous Acid

“…A mutant in the algH gene, which encodes a hypothetical protein that has been crystallized and predicted to be a potential redox sensor, was HOCl sensitive ( Fig. 4A ) ( 41 ). algH is part of a 4-gene operon that includes gshB , which encodes a glutathione synthetase; a gshB mutant was tested, and it displayed HOCl sensitivity ( Fig.…”

“…Catabolite repression by the Crc protein appears required for protection against HOCl, possibly indicating that optimal regulation of metabolic flexibility aids defense against HOCl. In step 2, the transcriptional regulator RclR, the H 2 O 2 sensor OxyR ( 42 ), and the hypothetical protein AlgH ( 41 ) all have putative redox-sensing capabilities and were identified as playing a role in protection against HOCl. In step 3, macromolecular repair mechanisms are implicated in the response to HOCl and HOSCN.…”

Response ofPseudomonas aeruginosato the innate immune system-derived oxidants hypochlorous acid and hypothiocyanous acid

“…A mutant in the algH gene, which encodes a hypothetical protein that has been crystallised and predicted to be a potential redox-sensor, was HOClsensitive ( Fig. 4A) (44). The algH gene is on a 4gene operon that includes gshB, which encodes a glutathione synthetase; a gshB mutant was tested and displayed HOCl-sensitivity ( Fig.…”