Dataset Information

Structural insights on the efficient catalysis of hydroperoxide reduction by Ohr: Crystallographic and molecular dynamics approaches.

ABSTRACT: Organic hydroperoxide resistance (Ohr) enzymes are highly efficient Cys-based peroxidases that play central roles in bacterial response to fatty acid hydroperoxides and peroxynitrite, two oxidants that are generated during host-pathogen interactions. In the active site of Ohr proteins, the conserved Arg (Arg19 in Ohr from Xylella fastidiosa) and Glu (Glu51 in Ohr from Xylella fastidiosa) residues, among other factors, are involved in the extremely high reactivity of the peroxidatic Cys (Cp) toward hydroperoxides. In the closed state, the thiolate of Cp is in close proximity to the guanidinium group of Arg19. Ohr enzymes can also assume an open state, where the loop containing the catalytic Arg is far away from Cp and Glu51. Here, we aimed to gain insights into the putative structural switches of the Ohr catalytic cycle. First, we describe the crystal structure of Ohr from Xylella fastidiosa (XfOhr) in the open state that, together with the previously described XfOhr structure in the closed state, may represent two snapshots along the coordinate of the enzyme-catalyzed reaction. These two structures were used for the experimental validation of molecular dynamics (MD) simulations. MD simulations employing distinct protonation states and in silico mutagenesis indicated that the polar interactions of Arg19 with Glu51 and Cp contributed to the stabilization of XfOhr in the closed state. Indeed, Cp oxidation to the disulfide state facilitated the switching of the Arg19 loop from the closed to the open state. In addition to the Arg19 loop, other portions of XfOhr displayed high mobility, such as a loop rich in Gly residues. In summary, we obtained a high correlation between crystallographic data, MD simulations and biochemical/enzymatic assays. The dynamics of the Ohr enzymes are unique among the Cys-based peroxidases, in which the active site Arg undergoes structural switches throughout the catalytic cycle, while Cp remains relatively static.

SUBMITTER: Piccirillo E

PROVIDER: S-EPMC5962072 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structural insights on the efficient catalysis of hydroperoxide reduction by Ohr: Crystallographic and molecular dynamics approaches.

Piccirillo Erika E Alegria Thiago G P TGP Discola Karen F KF Cussiol José R R JRR Domingos Renato M RM de Oliveira Marcos A MA Rezende Leandro de L Netto Luis E S LES Amaral Antonia T-do AT

PloS one 20180521 5

Organic hydroperoxide resistance (Ohr) enzymes are highly efficient Cys-based peroxidases that play central roles in bacterial response to fatty acid hydroperoxides and peroxynitrite, two oxidants that are generated during host-pathogen interactions. In the active site of Ohr proteins, the conserved Arg (Arg19 in Ohr from Xylella fastidiosa) and Glu (Glu51 in Ohr from Xylella fastidiosa) residues, among other factors, are involved in the extremely high reactivity of the peroxidatic Cys (Cp) towa ...[more]

PMID: 29782551

Similar Datasets

Project description:Hexachlorobenzene (HCB), as one of the persistent organic pollutants (POPs) and a possible human carcinogen, is especially resistant to biodegradation. In this study, HcbA1A3, a distinct flavin-N5-peroxide-utilizing enzyme and the sole known naturally occurring aerobic HCB dechlorinase, was biochemically characterized. Its apparent preference for HCB in binding affinity revealed that HcbA1 could oxidize only HCB rather than less-chlorinated benzenes such as pentachlorobenzene and tetrachlorobenzenes. In addition, the crystal structure of HcbA1 and its complex with flavin mononucleotide (FMN) were resolved, revealing HcbA1 to be a new member of the bacterial luciferase-like family. A much smaller substrate-binding pocket of HcbA1 than is seen with its close homologues suggests a requirement of limited space for catalysis. In the active center, Tyr362 and Asp315 are necessary in maintaining the normal conformation of HcbA1, while Arg311, Arg314, Phe10, Val59, and Met12 are pivotal for the substrate affinity. They are supposed to place HCB at a productive orientation through multiple interactions. His17, with its close contact with the site of oxidation of HCB, probably fixes the target chlorine atom and stabilizes reaction intermediates. The enzymatic characteristics and crystal structures reported here provide new insights into the substrate specificity and catalytic mechanism of HcbA1, which paves the way for its rational engineering and application in the bioremediation of HCB-polluted environments.IMPORTANCE As an endocrine disrupter and possible carcinogen to human beings, hexachlorobenzene (HCB) is especially resistant to biodegradation, largely due to difficulty in its dechlorination. The lack of knowledge of HCB dechlorinases limits their application in bioremediation. Recently, an HCB monooxygenase, HcbA1A3, representing the only naturally occurring aerobic HCB dechlorinase known so far, was reported. Here, we report its biochemical and structural characterization, providing new insights into its substrate selectivity and catalytic mechanism. This research also increases our understanding of HCB dechlorinases and flavin-N5-peroxide-utilizing enzymes.

Dataset Information

Structural insights on the efficient catalysis of hydroperoxide reduction by Ohr: Crystallographic and molecular dynamics approaches.

Publications

Structural insights on the efficient catalysis of hydroperoxide reduction by Ohr: Crystallographic and molecular dynamics approaches.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets