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Expression of the Caldariomyces fumago chloroperoxidase in Aspergillus niger and characterization of the recombinant enzyme

Author: Conesa, A. · Velde, F. van de · Rantwijk, F. van · Sheldon, R.A. · Hondel, C.A.M.J.J. van den · Punt, P.J.
Type:article
Date:2001
Institution: Centraal Instituut voor Voedingsonderzoek TNO
Source:Journal of Biological Chemistry, 21, 276, 17635-17640
Identifier: 72309
doi: doi:10.1074/jbc.M010571200
Keywords: Biology · Aspergillus · Aspergillus niger · Fungi · Leptoxyphium fumago · Caldariomyces · Aspergillus niger · Catalysis · Chloride Peroxidase · Fungal Proteins · Recombinant Proteins · Substrate Specificity

Abstract

The Caldariomyces fumago chloroperoxidase was successfully expressed in Aspergillus niger. The recombinant enzyme was produced in the culture medium as an active protein and could be purified by a three-step purification procedure. The catalytic behavior of recombinant chloroperoxidase (rCPO) was studied and compared with that of native CPO. The specific chlorination activity (47 units/nmol) of rCPO and its pH optimum (pH 2.75) were very similar to those of native CPO. rCPO catalyzes the oxidation of various substrates in comparable yields and selectivities to native CPO. Indole was oxidized to 2-oxindole with 99% selectivity and thioanisole to the corresponding R-sulfoxide (enantiomeric excess >98%). Incorporation of 18O from labeled H218O2 into the oxidized products was 100% in both cases. Molecular Sequence Numbers: GENBANK: AJ300448; Chemicals/CAS: chloride peroxidase, 9055-20-3; hydrogen, 12385-13-6, 1333-74-0; indole, 120-72-9; oxindole, 59-48-3; oxygen, 7782-44-7; sulfoxide, 120-62-7; thioanisole, 100-68-5; Chloride Peroxidase, EC 1.11.1.10; Fungal Proteins; Recombinant Proteins