用于制备酶生物反应器的一种转氨酶的纯化与固定(英文).pdfVIP

GE Healthcare Life Sciences Application note 29-0211-99 AA Chromatography systems Purification and immobilization of a transaminase for the preparation of an enzyme bioreactor A histidine-tagged transaminase was overexpressed in host and the crude extract of the material is used for mixing E. coli and purified using immobilized metal ion affinity with the precursor to be modified. The method is relatively chromatography (IMAC). Purification was achieved in a straightforward but has the drawback that purification of single step using HisTrap™ HP columns on ÄKTA™ pure the product becomes more complex due to the addition of chromatography system. Using the predefined NHS coupling large amounts of biomass without catalytic function. The use method in UNICORN™ v6.3 software for ÄKTA pure of purified enzyme reduces the purification efforts required system, the purified enzyme was immobilized on HiTrap™ to some extent. An even more interesting approach is to use NHS-activated HP columns for the preparation of an enzyme immobilized on a solid support for easy separation of enzyme bioreactor to enable catalysis of chiral amination product substance from the enzyme (1). Immobilization can reactions. Columns with immobilized transaminase were give increased stability of the enzyme, allowing the use of the used to investigate their efficiency in catalyzing conversion bioreactor for an extended period of time. of a model substrate applied to the column. The study, A proof-of-principle for the use of NHS-activated Sepharose which included use of the UNICORN Design of Experiment High Performance medium