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B6db references: 9531508

type Journal Article
authors Ravanel, S.; Gakiere, B.; Job, D.; Douce, R.
title Cystathionine gamma-synthase from Arabidopsis thaliana: purification and biochemical characterization of the recombinant enzyme overexpressed in Escherichia coli
journal Biochem J
Activity 2.5.1.48
Family 2.5.1.48.b
sel selected
ui 9531508
year (1998)
volume 331
number Pt 2
pages 639-48
 
keywords Alkynes/pharmacology
abstract Cystathionine gamma-synthase catalyses the first reaction specific for methionine biosynthesis in plants, the gamma-replacement of the phosphoryl substituent of O-phosphohomoserine by cysteine. A cDNA encoding cystathionine gamma-synthase from Arabidopsis thaliana has been cloned and used to overexpress the enzyme in Escherichia coli. The native recombinant enzyme is a homotetramer composed of 53 kDa subunits, each being tightly associated with one molecule of pyridoxal 5'-phosphate that binds at lysine-379 of the protein precursor. The replacement reaction follows a Ping Pong mechanism with a Vmax of 33.6 units/mg and Km values of 2.5 mM and 460 microM for O-phosphohomoserine and cysteine respectively. The protective effect of O-phosphohomoserine against enzyme inactivation by propargylglycine indicated that the Kd for the substrate is approx. 1/2500 of its Km value. Thus most of these biochemical properties are similar to those previously reported for plant and bacterial cystathionine gamma-synthases. However, the plant enzyme differs markedly from its enterobacterial counterparts because it catalyses a very faint gamma-elimination of O-phosphohomoserine in the absence of cysteine, this process being about 1/2700 as fast as the gamma-replacement reaction and approx. 1/1500 as fast as the gamma- elimination catalysed by the E. coli enzyme. This huge difference could be attributed to the inability of the A. thaliana cystathionine gamma- synthase to accumulate a long-wavelength-absorbing species that is characteristic for the efficient gamma-elimination reaction catalysed by the enterobacterial enzyme.
last changed 2009/02/11 18:24

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