Structural implications of the C-terminal tail in the catalytic and stability properties of manganese peroxidases from ligninolytic fungi

Fernández‐Fueyo, Elena; Acebes, Sandra; Ruíz-Dueñas, Francisco J.; Martı́nez, Marı́a Jesús; Romero, Antonio; Medrano, F.J.; Guallar, Vı́ctor; Martı́nez, Ángel T.

doi:10.1107/s1399004714022755

Cited by 34 publications

(54 citation statements)

References 40 publications

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“…As mentioned, lack of oxidation in MnP6 has been rationalized by the extra C-terminal residues, and ABTS oxidation in MnP4 has been attributed to the larger opening in the propionates 14 . The substrate migration, however, reveals the best minimum in MnP4 placed at the main heme channel ( Figure 1C, see also the supporting video), with a second minimum (~7 kcal/mol higher in energy) located at the propionate channel.…”

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“…The other one, known as the main heme channel, is located in the heme edge at the distal histidine side. At first sight, the main difference between MnP4 and MnP6 (with 46% sequence identity and 57% sequence similarity) is the C-terminal tail: 29 extra residues in MnP6 block the propionate channel ( Figure 1A), deletion of which introduces ABTS oxidation in this enzyme at the expense of thermostability decrease 14 .…”

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Rational Enzyme Engineering Through Biophysical and Biochemical Modeling

et al. 2016

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Due to its importance in the pharmaceutical industry, ligand dynamic simulations have experienced a great expansion. Using all-atom models and cutting edge hardware, one can perform non-biased ligand migration, active site search and binding studies. In this letter we demonstrate (and validate by PCR mutagenesis) how these techniques, when combined with quantum mechanics, open new possibilities in enzyme engineering. We provide a complete analysis where: 1) biophysical simulations produce ligand diffusion and, 2) biochemical modeling samples the chemical event. Using such broad analysis we engineer a highly stable peroxidase activating the enzyme for new substrate\ud oxidation after rational mutation of two non-conserved surface residues. In particular, we create a new surface-binding site, quantitatively predicting the in vitro change in oxidation rate obtained by mutagenic PCR and achieving a comparable specificity constant to active peroxidases.This work was supported by the INDOX (KBBE-2013-7-613549 to ATM) European project, and the CTQ2013-48287 (to VG) and BIO2014-56388-R (to FJR-D) projects of the Spanish Ministry of Economy and Competitiveness (MINECO). FJR-D acknowledges a MINECO\ud Ramón&Cajal contract.Peer ReviewedPostprint (author's final draft

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Rational Enzyme Engineering Through Biophysical and Biochemical Modeling

et al. 2016

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“…and to a major acidic stability, but also showed that only short MnPs have the capacity of oxidizing ABTS independently (in the absence of Mn 2+ ) [78]. Fernández-Fueyo et al using PELE [56] and electronic couplings calculations by the fragment charge difference (FCD) approach [80], provided computational information for the ABTS oxidation at the heme-propionate channel and the relation between the C-tail length and the MnPs catalytic properties.…”

Section: Manganese Peroxidasementioning

confidence: 99%

“…Also, the total energy during the molecular dynamics remained without major variations, confirming the stability of the system. The role of the C-terminal tail in the catalysis and stability of MnP from Ceriporiopsis subvermispora was studied through dynamic ligand diffusion and QM/MM techniques [78]. In a study of over 31 fungal genomes [79], 3 MnP subfamilies were defined depending on the length of the C-terminal tail: short, long and extralong MnPs.…”

Section: Manganese Peroxidasementioning

confidence: 99%

“…Since only small differences exist between long and extralong MnPs, there was no representative for the long MnP subfamily. Calculations were performed using the crystal structure of the extralong isoenzyme MnP6 [78] and its in silico shortened-tail form. Local exploration of the ABTS diffusion at the entrance of the heme-propionate channel using PELE showed that for the shortened-tail form of MnP6 the most favorable structures had ABTS closer to the active site.…”

Section: Manganese Peroxidasementioning

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Computational Modeling Methods for Understanding the Interaction of Lignin and Its Derivatives with Oxidoreductases as Biocatalysts

Alzate‐Morales¹,

Recabarren²,

Fuenzalida-Valdivia³

et al. 2018

Lignin - Trends and Applications

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This chapter will be presented as follow. First, a brief introduction to structure and characterization of lignin and its derivatives is presented, as well as their importance as chemical scaffolds for obtaining value-added products in chemical, food, pharmaceutical and agriculture industry. Second, an extensive review of different reports using computational modeling methods-like molecular dynamics simulations, quantum mechanics and hybrid calculation methods, among others-in the understanding of enzyme-substrate interaction and biocatalysis will be presented. Third, and as last part of chapter, some hand picked examples from literature will be chosen as successful cases where the interplay between experiment and computation has given as a result protein engineered oxidoreductases with improved catalytic capabilities.

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Redesign of a New Manganese Peroxidase Highly Expressed in Pichia pastoris towards a Lignin‐Degrading Versatile Peroxidase

2018

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Manganese peroxidases, lignin peroxidases, and versatile peroxidases secreted by white rot fungi are supposed to play an essential role in lignin degradation. Thus, these enzymes have attracted significant attention as potential biocatalysts. Versatile peroxidases are the most interesting ones, since they comprise activities of manganese and lignin peroxidases. Herein, we demonstrate how the properties of a new manganese peroxidase from Moniliophthora roreri, designated MrMnP1, were shifted towards those of a versatile peroxidase. MrMnP1 was cloned in Pichia pastoris X‐33 and highly expressed in a fed‐batch fermentation, yielding 132 mg L−1 of active enzyme. To extend the substrate spectrum of MrMnP1, a catalytically active tryptophan present in lignin and versatile peroxidases was first introduced. Additionally, the role of five amino acids at positions adjacent to the catalytic tryptophan was elucidated through their replacement by those found in a versatile peroxidase from Pleurotus eryngii. The resulting mutants demonstrated new activities towards high‐redox‐potential substrates, such as lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5.

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Structural implications of the C-terminal tail in the catalytic and stability properties of manganese peroxidases from ligninolytic fungi

Cited by 34 publications

References 40 publications

Rational Enzyme Engineering Through Biophysical and Biochemical Modeling

Rational Enzyme Engineering Through Biophysical and Biochemical Modeling

Computational Modeling Methods for Understanding the Interaction of Lignin and Its Derivatives with Oxidoreductases as Biocatalysts

Redesign of a New Manganese Peroxidase Highly Expressed in Pichia pastoris towards a Lignin‐Degrading Versatile Peroxidase

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