Purification and kinetic studies of recombinant gibberellin dioxygenases.

ABSTRACT:

Background

The 2-oxoglutarate-dependent dioxygenases (2ODDs) of gibberellin (GA) biosynthesis have a key role in the metabolism of a major plant hormone. The activity of recombinant GA 2ODDs from many species has been characterised in detail, however little information relates to enzyme purification. Native GA 2ODDs displayed lability during purification.

Results

Two GA 2ODDs were expressed in Escherichia coli and purified to homogeneity. The GA 2-oxidase from Pisum sativum L., PsGA2OX1, was expressed as a glutathione s-transferase (GST) fusion. It was purified in the three steps of affinity chromatography, GST removal and gel filtration. Highly pure PsGA2OX1 was obtained at a yield of 0.3 mg/g of cells. It displayed a Km of 0.024 microM and a Vmax of 4.4 pkat/mg toward [1be

SUBMITTER: Lester DR 

PROVIDER: S-EPMC1262730 | biostudies-literature | 2005 Sep

REPOSITORIES: biostudies-literature