Overproduction, purification, and characterization of recombinant aspartate semialdehyde dehydrogenase from Arabidopsis thaliana

Protein Expr Purif. 2002 Feb;24(1):99-104. doi: 10.1006/prep.2001.1538.

Abstract

In plant and microorganisms, aspartate semialdehyde dehydrogenase (ASDH) produces the branch point intermediate between the lysine and threonine/methionine pathways. In this study, we report the first cDNA cloning, purification, and characterization of a plant ASDH. The Arabidopsis thaliana ASDH is an homodimeric enzyme composed of subunits of 36 kDa. The plant enzyme exhibited a specific activity of 26 micromol NADPH oxidized min(-1) mg(-1) of protein with a K(M) value for NADPH of 92 microM. ASDH showed cooperative behavior for aspartyl phosphate with a K(0.5) value of 37 microM.

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis / enzymology*
  • Arabidopsis / genetics
  • Aspartate-Semialdehyde Dehydrogenase / chemistry
  • Aspartate-Semialdehyde Dehydrogenase / genetics*
  • Aspartate-Semialdehyde Dehydrogenase / isolation & purification
  • Cloning, Molecular*
  • DNA, Complementary
  • Escherichia coli / genetics
  • Gene Expression
  • Kinetics
  • Molecular Sequence Data
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sequence Analysis, Protein
  • Sequence Homology, Amino Acid
  • Solutions

Substances

  • DNA, Complementary
  • Recombinant Proteins
  • Solutions
  • Aspartate-Semialdehyde Dehydrogenase