title: Structural and functional characterization of ochratoxinase, a novel mycotoxin-degrading enzyme.
abstract: Ochratoxin, with ochratoxin A as the dominant form, is one of the five major mycotoxins most harmful to humans and animals. It is produced by Aspergillus and Penicillium species and occurs in a wide range of agricultural products. Detoxification of contaminated food is a challenging health issue. In the present paper we report the identification, characterization and crystal structure (at 2.2 Å) of a novel microbial ochratoxinase from Aspergillus niger. A putative amidase gene encoding a 480 amino acid polypeptide was cloned and homologously expressed in A. niger. The recombinant protein is N-terminally truncated, thermostable, has optimal activity at pH ~6 and 66°C, and is more efficient in ochratoxin A hydrolysis than carboxypeptidase A and Y, the two previously known enzymes capable of degrading this mycotoxin. The subunit of the homo-octameric enzyme folds into a two-domain structure characteristic of a metal dependent amidohydrolase, with a twisted TIM (triosephosphateisomerase)-barrel and a smaller β-sandwich domain. The active site contains an aspartate residue for acid-base catalysis, and a carboxylated lysine and four histidine residues for binding of a binuclear metal centre.
authors.forename: Doreen
authors.initials: D
authors.lastname: Dobritzsch
authors.forename: Huaming
authors.initials: H
authors.lastname: Wang
authors.affiliation: *Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-17177 Stockholm, Sweden.
authors.forename: Gunter
authors.initials: G
authors.lastname: Schneider
authors.forename: Shukun
authors.initials: S
authors.lastname: Yu
mesh_terms: D000581:Amidohydrolases / Q000302:isolation & purification / Q000378:metabolism*; D001234:Aspergillus niger / Q000201:enzymology; D020134:Catalytic Domain; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008958:Models, Molecular; D009793:Ochratoxins / Q000378:metabolism*; D010649:Phenylalanine / Q000031:analogs & derivatives / Q000378:metabolism; D011487:Protein Conformation; D013379:Substrate Specificity
chemical_list: D005656:Fungal Proteins; D009793:Ochratoxins; C025589:ochratoxin A; D010649:Phenylalanine; C067359:4-methylphenylalanine; D000581:Amidohydrolases; C000592582:ochratoxinase, Aspergillus niger