CYTOSOLIC BETA-CYANOALANINE SYNTHASE ACTIVITY ATTRIBUTED TO CYSTEINE SYNTHASES IN COCKLEBUR SEEDS - PURIFICATION AND CHARACTERIZATION OF CYTOSOLIC CYSTEINE SYNTHASES

The activity of beta-cyanoalanine synthase (CAS, EC 4.4.1.9) in cotyle dons of cocklebur seeds (Xanthium pennsylvanicum Wallr.) was detected both in the soluble and particulate fractions. The CBS activity of the soluble fraction (cytosolic CAS activity) was 10 times higher than th at of the particulate fraction. The CAS activity of the particulate fr action was confirmed to be localized in the mitochondria. Both enzymat ic activities were clearly separated by non-denaturing PAGE. The enzym e with cytosolic CAS activity has been extensively purified and separa ted into three different forms designated as cyt-1, cyt-2, and cyt-3. According to the SDS-PAGE analysis, the three enzymes are estimated to be a homodimer composed of 35-kDa subunits. The purified enzymes show ed CS activity. Partial amino acid sequences of cyt-1 were determined and had a high homology with cysteine synthases (CS, EC 4.2.99.8) from other plant sources. The catalytic action of the purified CSs in conv erting cyanide and cysteine into H2S and beta-cyanoalanine was confirm ed by the detection of significant (CN)-C-14 incorporation into beta-c yanoalanine. These results indicated that cytosolic CAS activity is du e to cytosolic CS and suggested that the CAS activity of CS is likely to be involved in cyanide metabolism in plant tissues.