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Stability mechanisms of a thermophilic laccase probed by molecular dynamics.


ABSTRACT: Laccases are highly stable, industrially important enzymes capable of oxidizing a large range of substrates. Causes for their stability are, as for other proteins, poorly understood. In this work, multiple-seed molecular dynamics (MD) was applied to a Trametes versicolor laccase in response to variable ionic strengths, temperatures, and glycosylation status. Near-physiological conditions provided excellent agreement with the crystal structure (average RMSD ?0.92 Å) and residual agreement with experimental B-factors. The persistence of backbone hydrogen bonds was identified as a key descriptor of structural response to environment, whereas solvent-accessibility, radius of gyration, and fluctuations were only locally relevant. Backbone hydrogen bonds decreased systematically with temperature in all simulations (?9 per 50 K), probing structural changes associated with enthalpy-entropy compensation. Approaching T opt (?350 K) from 300 K, this change correlated with a beginning "unzipping" of critical ?-sheets. 0 M ionic strength triggered partial denucleation of the C-terminal (known experimentally to be sensitive) at 400 K, suggesting a general salt stabilization effect. In contrast, F(-) (but not Cl(-)) specifically impaired secondary structure by formation of strong hydrogen bonds with backbone NH, providing a mechanism for experimentally observed small anion destabilization, potentially remedied by site-directed mutagenesis at critical intrusion sites. N-glycosylation was found to support structural integrity by increasing persistent backbone hydrogen bonds by ?4 across simulations, mainly via prevention of F(-) intrusion. Hydrogen-bond loss in distinct loop regions and ends of critical ?-sheets suggest potential strategies for laboratory optimization of these industrially important enzymes.

SUBMITTER: Christensen NJ 

PROVIDER: S-EPMC3639223 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Stability mechanisms of a thermophilic laccase probed by molecular dynamics.

Christensen Niels J NJ   Kepp Kasper P KP  

PloS one 20130429 4


Laccases are highly stable, industrially important enzymes capable of oxidizing a large range of substrates. Causes for their stability are, as for other proteins, poorly understood. In this work, multiple-seed molecular dynamics (MD) was applied to a Trametes versicolor laccase in response to variable ionic strengths, temperatures, and glycosylation status. Near-physiological conditions provided excellent agreement with the crystal structure (average RMSD ∼0.92 Å) and residual agreement with ex  ...[more]

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