Clinical Chemistry
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Clinical Chemistry 40: 688-704, 1994;
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Clinical Chemistry, Vol 40, 688-704, Copyright © 1994 by American Association for Clinical Chemistry

Enzymes in diagnostics: achievements and possibilities of recombinant DNA technology

E Kopetzki, K Lehnert and P Buckel
Boehringer Mannheim GmbH, Department of Genetics, Penzberg, Germany.

We discuss, from an industrial point of view, the scope and possibilities of recombinant DNA technology for "diagnostic enzyme" production and application. We describe the construction of enzyme- overproducing strains and show how to simplify downstream processing, increase product quality and process profitability, improve diagnostic enzyme properties, and adjust enzymes to harsh assay conditions. We also consider some safety and environmental aspects of enzyme production. Other aspects of diagnostic enzymes that we cover are the facilitation of enzyme purification by attachment of short amino acid tails, the introduction of tails or tags for site-specific conjugation or oriented immobilization, the construction of bi- or multifunctional enzymes, and the production of enzyme-based diagnostic tests as demonstrated by the homogeneous immunoassay system of CEDIA tests. We use as examples of diagnostic enzymes glucose-6-phosphate dehydrogenase (EC 1.1.1.49), glucose oxidase (EC 1.1.3.4), alkaline phosphatase (EC 3.1.3.1), alpha-glucosidase (EC 3.2.1.20), pyruvate oxidase (EC 1.2.3.3), creatinase (EC 3.5.3.3), and beta-galactosidase (EC 3.2.1.23).


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