Characterization of a phosphotriesterase-like lactonase from the hyperthermoacidophilic crenarchaeon Vulcanisaeta moutnovskia

Kallnik, Verena; Bunescu, Alina; Sayer, C.; Bräsen, Christopher; Wohlgemuth, Roland; Littlechild, Jennifer A.; Siebers, Bettina

doi:10.1016/j.jbiotec.2014.04.026

Cited by 26 publications

(20 citation statements)

References 40 publications

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“…All enantiomers of the two substrates GVL and γ-caprolactone were tested to determine stereoselectivity of the enzyme. Results indicated that while activity is seen with both isomers the enzyme seems to favour the D form of these substrates (Kallnik et al, 2014). The V. moutnovskia lactonase structure (Hiblot et al, 2015) shows a substrate binding pocket lined with hydrophobic amino acid side chains and located in the enzyme structure by a bound long chain fatty acid.…”

Section: Vulcanisaeta Mountnovskia Quorum Sensing Type Lactonasementioning

confidence: 99%

“…A BLAST sequence comparison using the archaeal Sulfolobus lactonases has identified a putative lactonase in the recently sequenced genome of Vulcanisaeta mountnovskia with 52 % sequence identity. In order to determine if the V. moutnovskia had PTE or PLL activity cloning and overexpression of the predicted phosphotriesterase was done in E.coli, which was followed by protein purification and biochemical characterisation (Kallnik et al, 2014). The substrate specificity of the lactonase from V. moutnovskia was evaluated using industrially relevant substrates.…”

Section: Vulcanisaeta Mountnovskia Quorum Sensing Type Lactonasementioning

confidence: 99%

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Discovering novel hydrolases from hot environments

Wohlgemuth

Littlechild

Monti

et al. 2018

Biotechnology Advances

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Novel hydrolases from hot and other extreme environments showing appropriate performance and/or novel functionalities, and new approaches for their systematic screening are of great interest for developing new processes, for improving safety, health and environment issues. Existing processes could benefit as well from their properties. The workflow, based on the HotZyme project, describes a multitude of technologies and their integration from discovery to application, providing new tools for discovering, identifying and characterizing more novel thermostable hydrolases with desired functions from hot terrestrial and marine environments. To this end, hot springs worldwide were mined, resulting in hundreds of environmental samples and thousands of enrichment cultures growing on polymeric substrates of industrial interest. Using high-throughput sequencing and bioinformatics, 15 hot spring metagenomes, as well as several sequenced isolate genomes and transcriptomes were obtained. To facilitate the discovery of novel hydrolases, the annotation platform Anastasia and a whole-cell bioreporter-based functional screening method were developed. Sequence-based screening and functional screening together resulted in about 100 potentially new hydrolases of which more than a dozen have been characterized comprehensively from a biochemical and structural perspective. The characterized hydrolases include thermostable carboxylesterases, enol lactonases, quorum sensing lactonases, gluconolactonases, epoxide hydrolases, and cellulases. Apart from these novel thermostable hydrolases, the project generated an enormous amount of samples and data, thereby allowing the future discovery of even more novel enzymes.

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Section: Vulcanisaeta Mountnovskia Quorum Sensing Type Lactonasementioning

confidence: 99%

Section: Vulcanisaeta Mountnovskia Quorum Sensing Type Lactonasementioning

confidence: 99%

Discovering novel hydrolases from hot environments

Wohlgemuth

Littlechild

Monti

et al. 2018

Biotechnology Advances

Self Cite

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“…A lactonase of this class has been identified, cloned, over-expressed and characterised [34] from Vulcanisaeta moutnovskia a hyperthermoacidophilic crenarchaeon isolated from a solfataric field in Kamchatka (Russia) [23, 55]. This lactonase was studied with view to its substrate specificity for biocatalytic applications of interest to the pharmaceutical industries.…”

Section: Lactonasesmentioning

confidence: 99%

“…No measurable activity was seen for mevalonolactone or δ-decalactone. The lactonase enzyme had increased activity towards the D form of these substrates [34]. …”

Section: Lactonasesmentioning

confidence: 99%

Improving the ‘tool box’ for robust industrial enzymes

Littlechild

2017

Journal of Industrial Microbiology and Biotechnology

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The speed of sequencing of microbial genomes and metagenomes is providing an ever increasing resource for the identification of new robust biocatalysts with industrial applications for many different aspects of industrial biotechnology. Using ‘natures catalysts’ provides a sustainable approach to chemical synthesis of fine chemicals, general chemicals such as surfactants and new consumer-based materials such as biodegradable plastics. This provides a sustainable and ‘green chemistry’ route to chemical synthesis which generates no toxic waste and is environmentally friendly. In addition, enzymes can play important roles in other applications such as carbon dioxide capture, breakdown of food and other waste streams to provide a route to the concept of a ‘circular economy’ where nothing is wasted. The use of improved bioinformatic approaches and the development of new rapid enzyme activity screening methodology can provide an endless resource for new robust industrial biocatalysts.This mini-review will discuss several recent case studies where industrial enzymes of ‘high priority’ have been identified and characterised. It will highlight specific hydrolase enzymes and recent case studies which have been carried out within our group in Exeter.

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“…PLL family enzymes are closely related to mesophilic bdPTEs, being classified in the same amidohydrolase superfamily. Several thermostable PLL enzymes have been identified from either hyperthermophilic archaea or thermophilic/extremophilic bacteria [23][24][25][26][27]. Their characteristic thermal stability made PLL enzymes one of the ideal candidates to apply for OP-degradation in practical decontamination issues.…”

Section: Introductionmentioning

confidence: 99%

Structural and Functional Characterization of New SsoPox Variant Points to the Dimer Interface as a Driver for the Increase in Promiscuous Paraoxonase Activity

Suzumoto¹,

Dim

Roviello

et al. 2020

IJMS

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Increasing attention is more and more directed toward the thermostable Phosphotriesterase-Like-Lactonase (PLL) family of enzymes, for the efficient and reliable decontamination of toxic nerve agents. In the present study, the DNA Staggered Extension Process (StEP) technique was utilized to obtain new variants of PLL enzymes. Divergent homologous genes encoding PLL enzymes were utilized as templates for gene recombination and yielded a new variant of SsoPox from Saccharolobus solfataricus. The new mutant, V82L/C258L/I261F/W263A (4Mut) exhibited catalytic efficiency of 1.6 × 10 5 M −1 s −1 against paraoxon hydrolysis at 70 • C, which is more than 3.5-fold and 42-fold improved in comparison with C258L/I261F/W263A (3Mut) and wild type SsoPox, respectively. 4Mut was also tested with chemical warfare nerve agents including tabun, sarin, soman, cyclosarin and VX. In particular, 4Mut showed about 10-fold enhancement in the hydrolysis of tabun and soman with respect to 3Mut. The crystal structure of 4Mut has been solved at the resolution of 2.8 Å. We propose that, reorganization of dimer conformation that led to increased central groove volume and dimer flexibility could be the major determinant for the improvement in hydrolytic activity in the 4Mut.

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Characterization of a phosphotriesterase-like lactonase from the hyperthermoacidophilic crenarchaeon Vulcanisaeta moutnovskia

Cited by 26 publications

References 40 publications

Discovering novel hydrolases from hot environments

Discovering novel hydrolases from hot environments

Improving the ‘tool box’ for robust industrial enzymes

Structural and Functional Characterization of New SsoPox Variant Points to the Dimer Interface as a Driver for the Increase in Promiscuous Paraoxonase Activity

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