Production, properties and application to biocatalysis of a novel extracellular alkaline phenol oxidase from the thermophilic fungus Scytalidium thermophilum.
Scytalidium thermophilum produces an extracellular phenol oxidase on glucose-containing medium. Certain phenolic acids, specifically gallic acid and tannic acid, induce the expression of the enzyme. Production at 45 degrees C in batch cultures is growth-associated and is enhanced in the presence of 160 microM CuSO4 x 5 H2O and 3 mM gallic acid. The highest enzyme activity is observed at pH 7.5 and 65 degrees C, on catechol. When incubated for 1 h at pH 7 and pH 8, 95% and 86% of the activity is retained. Thermostability decreases gradually from 40 degrees C to 80 degrees C. Estimated molecular mass is c. 83 kDa, and pI is acidic at c. 5.4. Substrate specificity and inhibition analysis in culture supernatants suggest that the enzyme has unique properties showing activity towards catechol; 3,4-dihydroxy-L-phenylalanine (L-DOPA); 4-amino-N, N-diethylaniline (ADA); p-hydroquinone; gallic acid; tannic acid and caffeic acid, and no activity towards L-tyrosine, guaiacol, 2,2'-azino-bis(3-ethyl-benzthiazoline-6-sulphonic acid) (ABTS) and syringaldazine. Inhibition is observed in the presence of salicyl hydroxamic acid (SHAM) and p-coumaric acid. Enzyme activity is enhanced by cetyltrimethylammonium bromide (CTAB) and polyvinylpyrrolidone (PVP), and the organic solvents dimethyl sulfoxide (DMSO) and ethanol. No inhibition is observed in the presence of carbon monoxide. Benzoin, benzoyl benzoin and hydrobenzoin are converted into benzil, and stereoselective oxidation is observed on hydrobenzoin. The reported enzyme is novel due to its catalytic properties resembling mainly catechol oxidases, but displaying some features of laccases at the same time.
Ogel
Z B
ZB
Food Engineering Dept., The Middle East Technical University, 06531, Ankara, Turkey. zogel@metu.edu.tr
Yüzügüllü
Y
Y
Mete
S
S
Bakir
U
U
Kaptan
Y
Y
Sutay
D
D
Demir
A S
AS
D001203:Ascomycota / Q000201:enzymology* / Q000378:metabolism; D001205:Ascorbic Acid / Q000494:pharmacology; D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003300:Copper / Q000494:pharmacology; D003373:Coumaric Acids / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D005707:Gallic Acid / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D015394:Molecular Structure; D008970:Molecular Weight; D014442:Monophenol Monooxygenase / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D010655:Phenylenediamines / Q000494:pharmacology; D012997:Solvents / Q000494:pharmacology; D013379:Substrate Specificity; D013696:Temperature; D013997:Time Factors
D003373:Coumaric Acids; D005656:Fungal Proteins; D010655:Phenylenediamines; D012997:Solvents; D005707:Gallic Acid; D003300:Copper; C013935:N,N-diethyl 4-phenylenediamine; D014442:Monophenol Monooxygenase; D001205:Ascorbic Acid