Catalytic chromatography

Catalytic chromatography exploits both specific biological affinity and cat alytic specificity to selectively purify enzymes. Two different application s are presented. Purification of EcoRI restriction endonuclease to apparent homogeneity was accomplished in a single step with significantly greater y ield and purification than was obtained with affinity chromatography. An at tempt to purify the multiple DNA polymerase activities of Escherichia coli was also developed. Five well-resolved peaks of DNA polymerase activity wer e fractionated. In this new chromatographic mode, the enzyme binds immobili zed substrate coupled to a column in the absence of some required cofactor. When the missing cofactor is added, the enzyme converts substrate to produ ct and selectively elutes from the column. (C) 2000 Academic Press.