Catalytic surface radical in dye-decolorizing peroxidase: a computational, spectroscopic and site-directed mutagenesis study
Dolores Linde,Rebecca Pogni,Marina Cañellas,Fátima Lucas,Victor Guallar,Maria Camilla Baratto,Adalgisa Sinicropi,Verónica Sáez-Jiménez,Cristina Coscolín,Antonio A. Romero,Francisco J. Medrano,Francisco J. Ruiz-Dueñas,Ángel T. Martínez +12 more
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Kinetics of substrate oxidation by DyP suggests the existence of high- and low-turnover sites, and QM/MM calculations showed a higher tendency of Trp-377 than other exposed haem-neighbouring residues to harbour a catalytic protein radical, and identified the electron-transfer pathway.Abstract:
Dye-decolorizing peroxidase (DyP) of Auricularia auricula-judae has been expressed in Escherichia coli as a representative of a new DyP family, and subjected to mutagenic, spectroscopic, crystallographic and computational studies. The crystal structure of DyP shows a buried haem cofactor, and surface tryptophan and tyrosine residues potentially involved in long-range electron transfer from bulky dyes. Simulations using PELE (Protein Energy Landscape Exploration) software provided several binding-energy optima for the anthraquinone-type RB19 (Reactive Blue 19) near the above aromatic residues and the haem access-channel. Subsequent QM/MM (quantum mechanics/molecular mechanics) calculations showed a higher tendency of Trp-377 than other exposed haem-neighbouring residues to harbour a catalytic protein radical, and identified the electron-transfer pathway. The existence of such a radical in H2O2-activated DyP was shown by low-temperature EPR, being identified as a mixed tryptophanyl/tyrosyl radical in multifrequency experiments. The signal was dominated by the Trp-377 neutral radical contribution, which disappeared in the W377S variant, and included a tyrosyl contribution assigned to Tyr-337 after analysing the W377S spectra. Kinetics of substrate oxidation by DyP suggests the existence of high- and low-turnover sites. The high-turnover site for oxidation of RB19 (kcat> 200 s−1) and other DyP substrates was assigned to Trp-377 since it was absent from the W377S variant. The low-turnover site/s (RB19 kcat ~20 s−1) could correspond to the haem access-channel, since activity was decreased when the haem channel was occluded by the G169L mutation. If a tyrosine residue is also involved, it will be different from Tyr-337 since all activities are largely unaffected in the Y337S variant.read more
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Oxidoreductases on their way to industrial biotransformations
Ángel T. Martínez,Francisco J. Ruiz-Dueñas,Susana Camarero,Ana Serrano,Dolores Linde,Henrik Lund,Jesper Vind,Morten Tovborg,Owik Matthias Herold-Majumdar,Martin Hofrichter,Christiane Liers,René Ullrich,Katrin Scheibner,Giovanni Sannia,Alessandra Piscitelli,Cinzia Pezzella,Mehmet E. Sener,Sibel Kilic,Willem J. H. van Berkel,Victor Guallar,María Lucas,Ralf Zuhse,Roland Ludwig,Frank Hollmann,Elena Fernández-Fueyo,Eric Record,Craig B. Faulds,Marta Tortajada,Ib Winckelmann,Jo-Anne Rasmussen,Mirjana Gelo-Pujic,Ana Gutiérrez,José C. del Río,Jorge Rencoret,Miguel Alcalde +34 more
TL;DR: The recently described lytic polysaccharide monooxygenases have attracted the highest attention among copper oxidoreductases, since they are capable of oxidatively breaking down crystalline cellulose, the disintegration of which is still a major bottleneck in lignocellulose biorefineries, along with lignin degradation.
Journal ArticleDOI
Characterization of Dye-decolorizing Peroxidase (DyP) from Thermomonospora curvata Reveals Unique Catalytic Properties of A-type DyPs
Chao Chen,Ruben Shrestha,Kaimin Jia,Philip Gao,Brian V. Geisbrecht,Stefan H. Bossmann,Jishu Shi,Ping Li +7 more
TL;DR: Results reveal the unique catalytic property of the A-type DyPs, which will facilitate the development of these enzymes as lignin degraders and allow development of biocatalysts for lign in degradation.
Journal ArticleDOI
Synthesis of 1-Naphthol by a Natural Peroxygenase Engineered by Directed Evolution
Patricia Molina-Espeja,Marina Cañellas,Francisco J. Plou,Martin Hofrichter,Fátima Lucas,Victor Guallar,Victor Guallar,Miguel Alcalde +7 more
TL;DR: Powered exclusively by catalytic concentrations of H2O2, this soluble and stable biocatalyst has a total turnover number of 50 000, with high regioselectivity (97 %) and reduced peroxidative activity.
Journal ArticleDOI
The multihued palette of dye-decolorizing peroxidases.
Rahul Singh,Lindsay D. Eltis +1 more
TL;DR: The phylogenetic analysis reveals the existence of at least five classes of DyPs, and their potential physiological roles are discussed based in part on synteny analyses and the considerable biotechnological potential is summarized.
Journal ArticleDOI
Basidiomycete DyPs: Genomic diversity, structural–functional aspects, reaction mechanism and environmental significance
Dolores Linde,Francisco J. Ruiz-Dueñas,Elena Fernández-Fueyo,Victor Guallar,Kenneth E. Hammel,Rebecca Pogni,Ángel T. Martínez +6 more
TL;DR: DyP degradation of substituted anthraquinone dyes most probably proceeds via typical one-electron peroxidase oxidations and product breakdown without a DyP-catalyzed hydrolase reaction, and a significant contribution to lignin degradation is unlikely because of the low activity on high redox-potential substrates.
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