Comparative spectroelectrochemical studies of lyophilized and nonlyophilized laccases from Cerrena unicolor basidiomycete

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Abstract

The electrochemical, spectroelectrochemical, and kinetic investigations of two preparations of Cerrena unicolor laccase, lyophilized (LLAC) and nonlyophilized frozen enzymes (FLAC), were performed. It was found that the value of the redox potential of the T1 site of C. unicolor laccase is ca. 750 vs. NHE. It was also shown that one of the redox potentials of the T2/T3 cluster of C. unicolor laccase is close to 400 mV, as was previously confirmed for other blue multicopper oxidases, such as trees and fungal laccases, ascorbate oxidase, and bilirubin oxidase. Furthermore, the poor stability of both preparations, but especially of LLAC, in their reduced state was confirmed using mediated and mediatorless spectroelectrochemical studies. DET-based biocatalytic reduction of O-2 by C unicolor laccase was only obtained, when FLAC was directly adsorbed on a spectrographic graphite electrode. Moreover, only low values of the steady-state potentials of gold and graphite electrodes modified by C. unicolor laccase were also found. Heterogeneity of the 3-D structures of laccase molecules, conformational changes, and partial denaturation of the enzyme, which appeared after enzyme isolation, purification, and especially lyophilization, were found to be the reasons for the low bioelectrocatalytic current, the high Km-value towards O-2, and the unusual electrochemical behavior of C unicolor laccase used in the present study. In spite of the comparable specific activity and long-term stability of both preparations in homogeneous solution, the stability of immobilized LLAC was found to be inadmissibly low for both fundamental studies and possible electrochemical applications. Indeed, FLAC is a much better source of enzyme than its lyophilized counterpart.

Detaljer

Författare
  • Sergey Shleev
  • Maciej Klis
  • Yan Wang
  • Jerzy Rogalski
  • Renata Bilewicz
  • Lo Gorton
Enheter & grupper
Forskningsområden

Ämnesklassifikation (UKÄ) – OBLIGATORISK

  • Analytisk kemi

Nyckelord

Originalspråkengelska
Sidor (från-till)1039-1047
TidskriftElectroanalysis
Volym19
Utgåva nummer10
StatusPublished - 2007
PublikationskategoriForskning
Peer review utfördJa