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Assembly of preactivation complex for urease maturation in Helicobacter pylori: crystal structure of UreF-UreH protein complex.

ABSTRACT: Colonization of Helicobacter pylori in the acidic environment of the human stomach depends on the neutralizing activity of urease. Activation of apo-urease involves carboxylation of lysine 219 and insertion of two nickel ions. In H. pylori, this maturation process involves four urease accessory proteins as follows: UreE, UreF, UreG, and UreH. It is postulated that the apo-urease interacts with UreF, UreG, and UreH to form a pre-activation complex that undergoes GTP-dependent activation of urease. The crystal structure of the UreF-UreH complex reveals conformational changes in two distinct regions of UreF upon complex formation. First, the flexible C-terminal residues of UreF become ordered, forming an extra helix ?10 and a loop structure stabilized by hydrogen bonds involving Arg-250. Second, the first turn of helix ?2 uncoils to expose a conserved residue, Tyr-48. Substitution of R250A or Y48A in UreF abolishes the formation of the heterotrimeric complex of UreG-UreF-UreH and abolishes urease maturation. Our results suggest that the C-terminal residues and helix ?2 of UreF are essential for the recruitment of UreG for the formation of the pre-activation complex. The molecular mass of the UreF-UreH complex determined by static light scattering was 116 ± 2.3 kDa, which is consistent with the quaternary structure of a dimer of heterodimers observed in the crystal structure. Taking advantage of the unique 2-fold symmetry observed in both the crystal structures of H. pylori urease and the UreF-UreH complex, we proposed a topology model of the pre-activation complex for urease maturation.

SUBMITTER: Fong YH

PROVIDER: S-EPMC3234868 | biostudies-literature | 2011 Dec

REPOSITORIES: biostudies-literature

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Assembly of preactivation complex for urease maturation in Helicobacter pylori: crystal structure of UreF-UreH protein complex.

Fong Yu Hang YH Wong Ho Chun HC Chuck Chi Pang CP Chen Yu Wai YW Sun Hongzhe H Wong Kam-Bo KB

The Journal of biological chemistry 20111019 50

Colonization of Helicobacter pylori in the acidic environment of the human stomach depends on the neutralizing activity of urease. Activation of apo-urease involves carboxylation of lysine 219 and insertion of two nickel ions. In H. pylori, this maturation process involves four urease accessory proteins as follows: UreE, UreF, UreG, and UreH. It is postulated that the apo-urease interacts with UreF, UreG, and UreH to form a pre-activation complex that undergoes GTP-dependent activation of urease ...[more]

PMID: 22013070

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Project description:The well-studied catalytic role of urease, the Ni-dependent conversion of urea into carbon dioxide and ammonia, has been shown to protect Helicobacter pylori against the low pH environment of the stomach lumen. We hypothesized that the abundantly expressed urease protein can play another noncatalytic role in combating oxidative stress via Met residue-mediated quenching of harmful oxidants. Three catalytically inactive urease mutant strains were constructed by single substitutions of Ni binding residues. The mutant versions synthesize normal levels of urease, and the altered versions retained all methionine residues. The three site-directed urease mutants were able to better withstand a hypochlorous acid (HOCl) challenge than a ?ureAB deletion strain. The capacity of purified urease to protect whole cells via oxidant quenching was assessed by adding urease enzyme to nongrowing HOCl-exposed cells. No wild-type cells were recovered with oxidant alone, whereas urease addition significantly aided viability. These results suggest that urease can protect H. pylori against oxidative damage and that the protective ability is distinct from the well-characterized catalytic role. To determine the capability of methionine sulfoxide reductase (Msr) to reduce oxidized Met residues in urease, purified H. pylori urease was exposed to HOCl and a previously described Msr peptide repair mixture was added. Of the 25 methionine residues in urease, 11 were subject to both oxidation and to Msr-mediated repair, as identified by mass spectrometry (MS) analysis; therefore, the oxidant-quenchable Met pool comprising urease can be recycled by the Msr repair system. Noncatalytic urease appears to play an important role in oxidant protection.IMPORTANCE Chronic Helicobacter pylori infection can lead to gastric ulcers and gastric cancers. The enzyme urease contributes to the survival of the bacterium in the harsh environment of the stomach by increasing the local pH. In addition to combating acid, H. pylori must survive host-produced reactive oxygen species to persist in the gastric mucosa. We describe a cyclic amino acid-based antioxidant role of urease, whereby oxidized methionine residues can be recycled by methionine sulfoxide reductase to again quench oxidants. This work expands our understanding of the role of an already acknowledged pathogen virulence factor and specifically expands our knowledge of H. pylori survival mechanisms.

Dataset Information

Assembly of preactivation complex for urease maturation in Helicobacter pylori: crystal structure of UreF-UreH protein complex.

Publications

Assembly of preactivation complex for urease maturation in Helicobacter pylori: crystal structure of UreF-UreH protein complex.

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