Kinetic and redox properties of MnP II, a major manganese peroxidase isoenzyme from Panus tigrinus CBS 577.79.
A manganese peroxidase (MnP) isoenzyme from Panus tigrinus CBS 577.79 was produced in a benchtop stirred-tank reactor and purified to apparent homogeneity. The purification scheme involving ultrafiltration, affinity chromatography on concanavalin-A Sepharose, and gel filtration led to a purified MnP, termed "MnP II," with a specific activity of 288 IU mg(-1) protein and a final yield of 22%. The enzyme turned out to be a monomeric protein with molecular mass of 50.5 kDa, pI of 4.07, and an extent of N-glycosylation of about 5.3% of the high-mannose type. The temperature and pH optima for the formation of malonate manganic chelates were 45 degrees C and 5.5, respectively. MnP II proved to be poorly thermostable at 50 and 60 degrees C, with half-lives of 11 min and 105 s, respectively. K (m) values for H(2)O(2) and Mn(2+) were 16 and 124 microM, respectively. Although MnP II was able to oxidize veratryl alcohol and to catalyze the Mn(2+)-independent oxidation of several phenols, it cannot be assigned to the versatile peroxidase family. As opposed to versatile peroxidase oxidation, veratryl alcohol oxidation required the simultaneous presence of H(2)O(2) and Mn(2+); in addition, low turnover numbers and K (m) values higher than 300 microM characterized the Mn(2+)-independent oxidation of substituted phenols. Kinetic properties and the substrate specificity of the enzyme markedly differed from those reported for MnP isoenzymes produced by the reference strain P. tigrinus 8/18. To our knowledge, this study reports for the first time a thorough electrochemical characterization of a MnP from this fungus.
Petruccioli
Maurizio
M
Dipartimento di Agrobiologia and Agrochimica, Università degli Studi della Tuscia, 01100 Viterbo, Italy.
Frasconi
Marco
M
Quaratino
Daniele
D
Covino
Stefano
S
Favero
Gabriele
G
Mazzei
Franco
F
Federici
Federico
F
D'Annibale
Alessandro
A
D055664:Electrochemical Techniques; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020074:Lentinula / Q000201:enzymology*; D008345:Manganese / Q000378:metabolism; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D013696:Temperature
D005656:Fungal Proteins; D007527:Isoenzymes; D008345:Manganese; D010544:Peroxidases; C051129:manganese peroxidase