Separation, subcellular location and influence of sulphur nutrition on isoforms of cysteine synthase in spinach

Cysteine synthase (O-acetylserine (thiol) lyase, EC4.2.99.8) and -cyanoalanine synthase (EC4.4.1.9) have been isolated from leaves of Spinaceaoleracea L. and separated by anion-exchange chromatography.Further separation of one minor and two major isoforms of cysteine synthasewas achieved by hydrophobic interaction chromatography and high resolutionnative electrophoresis (PAGE). Analysis of root material indicated thatamongst the multiple isoforms present, a single isoform predominated.Subcellular fractionation studies indicated that one of the major leafforms, cysteine synthase B, was located in the chloroplast and the other,cysteine synthase B, occurred in the cytoplasm. No specific isoform ofcysteine synthase was resolved in the mitochondria, while cyanoalanine synthase was predominantly located in the mitochondrial fraction. Sulphurdeprivation decreased cysteine synthase activity, but not cyanalaninesynthase activity in both young and mature leaves, although cysteinesynthase activity in the roots increased slightly. A selective decrease incystein synthase B (chloroplastic abundance was observed in mature leaves.Patterns of expression of cysteine synthase in response to S-availabilityare discussed in relation to possible roles for this enzyme in controllingS-flux through the S-assimilatory pathway.Key words: Cysteine synthaseisoforms (expression of), Spinacea oleracea L.,sulphur deficiency.