IMPROVING ENZYME-ELECTRODE CONTACTS BY REDOX MODIFICATION OF COFACTORS

EFFICIENT electron transfer of redox proteins to and from their enviro nment is essential for the use of such proteins in biotechnological ap plications such as amperometric biosensors and photosynthetic biocatal ysts(1-3). But most redox enzymes lack pathways that can transport an electron from their embedded redox site to an electrode(4,5) or a diff using photoexcited species(6). Electrical communication between redox proteins and electrode surfaces has been improved by aligning proteins on chemically modified electrodes(7-9), by attaching electron-transpo rting groups(10,11) and by immobilizing proteins in polymer matrices t ethered by redox groups(12-14). Generally these methods involve contac ting the enzymes at random with electron relay units, Here we report a n approach that allows site-specific positioning of electron-mediating units in redox proteins, We strip glucose oxidase of its flavin adeni ne dinucleotide (FAD) cofactors, modify tbe latter with redox-active f errocene-containing groups, and then reconstitute the apoprotein with these modified cofactors, In this way, electrical contact between an e lectrode and the resulting enzyme in solution is greatly enhanced in a controlled and reproducible way.