IMMOBILIZATION OF MANGANESE PEROXIDASE FROM LENTINULA-EDODES ON ALKYLAMINATED EMPHAZE(TM) AB-1 POLYMER FOR GENERATION OF MN3-AGENT( AS AN OXIDIZING)

Manganese peroxidase (MnP) is secreted by white-rot fungi and particip ates in the degradation of lignin by these organisms. MnP uses H2O2 as an oxidant to oxidize Mn-II to Mn-III as the manganic ion Mn3+. The M n3+ stabilized by chelation, is a highly reactive nonspecific oxidant capable of oxidizing a variety of toxic organic compounds. Previous at tempts at immobilization of MnP, purified from Lentinula edodes throug h reactive amino groups, have been hindered by the protein's low lysin e content of only 1% and its instability above pH 6.0. As an alternati ve to amine coupling, the enzyme has now been covalently immobilized t hrough its carboxyl groups, using an azlactone-functional copolymer de rivatized with ethylenediamine and 2-ethoxy-1-ethoxycarbonyl-1,2-dihyd roquinoline (EEDQ) as a coupling reagent. The immobilization reaction was performed under acidic (pH 5.25) conditions, and 90% coupling effi ciency was achieved within 2 h. Net immobilization efficiencies, expre ssed as the product of protein coupling efficiency and enzyme activity , have been measured at > 95% within 4 h. The MnP-NH-polymer and the f ree soluble protein were characterized and compared for their pH, temp erature, and storage stabilities, as well as their H2O2 dependence and kinetics. The tethered MnP, employed in an immobilized enzyme bioreac tor for generation of chelated Mn3+ may have industrial applications a s a nonspecific oxidant of organopollutants.