Publication

Article Metrics

Citations


Online attention

A novel bacterial class V dye-decolourizing peroxidase from the extremophile Deinococcus radiodurans : cloning, expression optimization, purification, crystallization, initial characterization and X-ray diffraction analysis

DOI: 10.1107/S2053230X18008488 DOI Help

Authors: Kelly Stefany Tuna Frade (Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa (ITQB-NOVA)) , Andreia Cecília Pimenta Fernandes (Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa (ITQB-NOVA)) , Celia Marisa Silveira (Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa (ITQB-NOVA)) , Carlos Frazao (Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa (ITQB-NOVA)) , Elin Moe (Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa (ITQB-NOVA))
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Acta Crystallographica Section F Structural Biology Communications , VOL 74 , PAGES 419 - 424

State: Published (Approved)
Published: July 2018

Abstract: Deinococcus radiodurans is a bacterium with extreme resistance to desiccation and radiation. The resistance mechanism is unknown, but an efficient reactive oxygen species (ROS) scavenging system and DNA-repair and DNA-protection mechanisms are believed to play important roles. Here, the cloning and small- and medium-scale expression tests of a novel dye-decolourizing peroxidase from D. radiodurans (DrDyP) using three different Escherichia coli strains and three different temperatures in order to identify the optimum conditions for the expression of recombinant DrDyP are presented. The best expression conditions were used for large-scale expression and yielded ∼10 mg recombinant DrDyP per litre of culture after purification. Initial characterization experiments demonstrated unusual features with regard to the haem spin state, which motivated the crystallization experiment. The obtained crystals were used for data collection and diffracted to 2.2 Å resolution. The crystals belonged to the trigonal space group P31 or P32, with unit-cell parameters a = b = 64.13, c = 111.32 Å, and are predicted to contain one DrDyP molecule per asymmetric unit. Structure determination by molecular replacement using previously determined structures of dye-decolourizing peroxidases with ∼30% sequence identity at ∼2 Å resolution as templates are ongoing.

Journal Keywords: peroxidases; oxidative stress; radiation resistance; dye-decolourizing peroxidases; Deinococcus radiodurans

Subject Areas: Biology and Bio-materials


Instruments: I03-Macromolecular Crystallography

Other Facilities: ESRF