Resolution of the phase-ambiguity problem in the centrosymmetric P \bar{1} space group by Monte Carlo methods

Delarue, Marc

doi:10.1107/s0108767300008849

Cited by 2 publications

(1 citation statement)

References 38 publications

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“…The 2-fold noncrystallographic symmetry (NCS) operator was found using the program RESOLVE (42). Density modification, phase extension, and NCS averaging were performed using the program DM (43).…”

Section: Methodsmentioning

confidence: 99%

Pyrethroid Carboxylesterase PytH from Sphingobium faniae JZ-2: Structure and Catalytic Mechanism

Gao

Sun

et al. 2020

Appl Environ Microbiol

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Carboxylesterase PytH, isolated from the pyrethroid-degrading bacterium Sphingobium faniae JZ-2, could rapidly hydrolyze the ester bond of a wide range of pyrethroid pesticides, including permethrin, fenpropathrin, cypermethrin, fenvalerate, deltamethrin, cyhalothrin, and bifenthrin. To elucidate the catalytic mechanism of PytH, we report here the crystal structures of PytH with bifenthrin (BIF) and phenylmethylsulfonyl fluoride (PMSF) and two PytH mutants. Though PytH shares low sequence identity with reported α/β-hydrolase fold proteins, the typical triad catalytic center with Ser-His-Asp triad (Ser78, His230, and Asp202) is present and vital for the hydrolase activity. However, no contact was found between Ser78 and His230 in the structures we solved, which may be due to the fact that the PytH structures we determined are in their inactive or low-activity forms. The structure of PytH is composed of a core domain and a lid domain; some hydrophobic amino acid residues surrounding the substrate from both domains form a deeper and wider hydrophobic pocket than its homologous structures. This indicates that the larger hydrophobic pocket makes PytH fit for its larger substrate binding; both lid and core domains are involved in substrate binding, and the lid domain-induced core domain movement may make the active center correctly positioned with substrates. IMPORTANCE Pyrethroid pesticides are widely applied in agriculture and household; however, extensive use of these pesticides also causes serious environmental and health problems. The hydrolysis of pyrethroids by carboxylesterases is the major pathway of microbial degradation of pyrethroids, but the structure of carboxylesterases and its catalytic mechanism are still unknown. Carboxylesterase PytH from Sphingobium faniae JZ-2 could effectively hydrolyze a wide range of pyrethroid pesticides. The crystal structures of PytH are solved in this study. This showed that PytH belongs to the α/β-hydrolase fold proteins with typical catalytic Ser-His-Asp triad, though PytH has a low sequence identity (about 20%) with them. The special large hydrophobic binding pocket enabled PytH to bind bigger pyrethroid family substrates. Our structures shed light on the substrate selectivity and the future application of PytH and deepen our understanding of α/β-hydrolase members.

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Section: Methodsmentioning

confidence: 99%

Pyrethroid Carboxylesterase PytH from Sphingobium faniae JZ-2: Structure and Catalytic Mechanism

Gao

Sun

et al. 2020

Appl Environ Microbiol

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General formalism for phase combination and phase refinement: a statistical thermodynamics approach in reciprocal space

Delarue

Orland

2000

Acta Crystallogr A Found Crystallogr

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The mean‐field optimization methodology has been used to recast in a single formalism the problem of phase optimization using an arbitrary energy function in the presence of an experimentally determined phase probability distribution function. It results naturally in the generalization of the notions of figure of merit and centroid phase where the weight of the energy refinement is controlled by an effective temperature in a self‐consistent manner. In the limit of high temperature, the formalism reduces of course to the Blow & Crick [Acta Cryst. (1959), 12, 794–802] classical treatment. If a model is available, Sim's [Acta Cryst. (1960), 13, 511–512] weighting scheme for a combined map appears as the first step of a refinement to be conducted until self‐consistency is achieved. Assuming that MIR phases exist and that they agree reasonably well with the phases of the model, a first‐order expansion gives an estimate of the changes of weights and phases to be performed for the Fourier synthesis. This provides for a new way of doing phase combination that might prove useful in challenging cases of model refinement, e.g. in large macromolecular complexes. Thermodynamic considerations have been used to discuss the best determination of weights in phase refinement; they also suggest that a variational expression of maximum likelihood is best suited as a target for refinement because it is the free energy of the system. The formalism readily allows use of solvent flattening, density averaging and the atomicity criterion to refine phases, and automatically assigns a figure of merit to each reflection. Numerical tests of the method are presented in an attempt to resolve the phase‐ambiguity problem of protein crystallography in the centrosymmetric P space group using an energy derived from the Sayre equation.

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Resolution of the phase-ambiguity problem in the centrosymmetric P \bar{1} space group by Monte Carlo methods

Cited by 2 publications

References 38 publications

Pyrethroid Carboxylesterase PytH from Sphingobium faniae JZ-2: Structure and Catalytic Mechanism

Pyrethroid Carboxylesterase PytH from Sphingobium faniae JZ-2: Structure and Catalytic Mechanism

General formalism for phase combination and phase refinement: a statistical thermodynamics approach in reciprocal space

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