Immobilization of bacterial luciferase and FMN reductase on glass rods.

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Abstract
Bacterial luciferase and NAD(P)H:FMN oxidoreductase isolated from Beneckea harveyi were covalently linked via diazotization to arylamine porous glass beads which was cemented onto plain glass rods. These immobilized enzymes are individually active and function to produce light via a coupled reaction utilizing NADH or NADPH. These enzymes have properties similar to the soluble forms with regard to pH and substrate optima and exhibit linearity in peak intensity of the initial flash of light emitted as a function of NADH or NADPH concentration. Linearity with NADH is obtained in the range of 1 pmol-50 nmol, and between 10 pmol-200 nmol for NADPH. The bound enzymes are stable and reusable. This immobilized system offers a rapid and inexpensive method of assaying low concentrations of NADH and NADPH and serves as a model to study the association of these 2 enzymes.