Immobilization of manganese peroxidase from Lentinula edodes on alkylaminated Emphaze AB 1 polymer for generation of Mn3+ as an oxidizing agent.
Manganese peroxidase (MnP) is secreted by white-rot fungi and participates in the degradation of lignin by these organisms. MnP uses H2O2 as an oxidant to oxidize MnII to MnIII as the manganic ion Mn3+. The Mn3+ stabilized by chelation, is a highly reactive nonspecific oxidant capable of oxidizing a variety of toxic organic compounds. Previous attempts at immobilization of MnP, purified from Lentinula edodes through reactive amino groups, have been hindered by the protein's low lysing content of only 1% and its instability above pH 6.0. As an alternative to amine coupling, the enzyme has now been covalently immobilized through its carboxyl groups, using an azlactone-functional copolymer derivatized with ethylenediamine and 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline (EEDQ) as a coupling reagent. The immobilization reaction was performed under acidic (pH 5.25) conditions, and 90% coupling efficiency was achieved within 2h. Net immobilization efficiencies, expressed as the product of protein coupling efficiency and enzyme activity, have been measured at > 95% within 4h. The MnP-NH-polymer and the free soluble protein were characterized and compared for their pH, temperature, and storage stabilities, as well as their H2O2 dependence and kinetics. The tethered MnP, employed in an immobilized enzyme bioreactor for generation of chelated Mn3+ may have industrial applications as a nonspecific oxidant of organopollutants.
Grabski
A C
AC
Bioseparations Facility, University of Wisconsin Biotechnology Center, Madison 53706, USA.
Rasmussen
J K
JK
Coleman
P L
PL
Burgess
R R
RR
D000363:Agaricales / Q000201:enzymology*; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D007700:Kinetics; D010544:Peroxidases / Q000378:metabolism*; D011108:Polymers; D011804:Quinolines; D013816:Thermodynamics
D004800:Enzymes, Immobilized; D007202:Indicators and Reagents; D011108:Polymers; D011804:Quinolines; C000470:EEDQ; D006861:Hydrogen Peroxide; D010544:Peroxidases; C051129:manganese peroxidase