title: A kinetic comparison of asparagine synthetase isozymes from higher plants.
abstract: Four previously identified maize asparagine synthetase (AsnS) genes and a soy AsnS gene have been cloned and expressed in Escherichia coli. The enzymes have been purified and kinetically characterized. The plant AsnS proteins were expressed mainly in the inclusion bodies although small amounts of one form (ZmAsnS2) were recovered in the soluble fraction. In order to measure the kinetic properties of these enzymes a sensitive assay based on the detection of Asn by HPLC has been developed. In addition a method to refold the recombinant plant AsnS to produce active enzyme has been developed. The plant AsnS enzymes are kinetically distinct with substantial differences in K(m) (Gln) and V(max) values when compared to each other. These differences may be important factors for transgenic studies using AsnS genes for crop improvement.
authors.affiliation: Monsanto Company, 700 Chesterfield Parkway West, St. Louis, MO 63198, USA. stephen.m.duff@monsanto.com
authors.forename: Stephen M G
authors.initials: SM
authors.lastname: Duff
authors.forename: Qungang
authors.initials: Q
authors.lastname: Qi
authors.forename: Timothy
authors.initials: T
authors.lastname: Reich
authors.forename: Xiaoyun
authors.initials: X
authors.lastname: Wu
authors.forename: Todd
authors.initials: T
authors.lastname: Brown
authors.forename: James H
authors.initials: JH
authors.lastname: Crowley
authors.forename: Brad
authors.initials: B
authors.lastname: Fabbri
mesh_terms: D001217:Aspartate-Ammonia Ligase / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics*; D002851:Chromatography, High Pressure Liquid; D002479:Inclusion Bodies; D007527:Isoenzymes / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000493:pharmacokinetics; D012995:Solubility; D013025:Glycine max / Q000201:enzymology*; D003313:Zea mays / Q000201:enzymology*
chemical_list: D007527:Isoenzymes; D010940:Plant Proteins; D011994:Recombinant Proteins; D001217:Aspartate-Ammonia Ligase