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

Shleev, Sergey LU ; Klis, Maciej ; Wang, Yan ; Rogalski, Jerzy ; Bilewicz, Renata and Gorton, Lo LU (2007) In Electroanalysis 19(10). p.1039-1047
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.... (More)
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. (Less)
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author
; ; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
T2, T1, redox potential, lyophilisation, laccase, Cerrena unicolor, T3, sites
in
Electroanalysis
volume
19
issue
10
pages
1039 - 1047
publisher
John Wiley & Sons Inc.
external identifiers
  • wos:000246836400003
  • scopus:34547595517
ISSN
1040-0397
DOI
10.1002/elan.200603841
language
English
LU publication?
yes
additional info
The information about affiliations in this record was updated in December 2015. The record was previously connected to the following departments: Analytical Chemistry (S/LTH) (011001004)
id
0bcde8b2-fa10-4d92-b7c9-9356dad2bd3f (old id 657871)
date added to LUP
2016-04-01 16:30:58
date last changed
2022-03-22 19:15:22
@article{0bcde8b2-fa10-4d92-b7c9-9356dad2bd3f,
  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.}},
  author       = {{Shleev, Sergey and Klis, Maciej and Wang, Yan and Rogalski, Jerzy and Bilewicz, Renata and Gorton, Lo}},
  issn         = {{1040-0397}},
  keywords     = {{T2; T1; redox potential; lyophilisation; laccase; Cerrena unicolor; T3; sites}},
  language     = {{eng}},
  number       = {{10}},
  pages        = {{1039--1047}},
  publisher    = {{John Wiley & Sons Inc.}},
  series       = {{Electroanalysis}},
  title        = {{Comparative spectroelectrochemical studies of lyophilized and nonlyophilized laccases from Cerrena unicolor basidiomycete}},
  url          = {{http://dx.doi.org/10.1002/elan.200603841}},
  doi          = {{10.1002/elan.200603841}},
  volume       = {{19}},
  year         = {{2007}},
}