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 cotyledons of cocklebur seeds (Xanthium pennsylvanicum Wallr.) was detected both in the soluble and particulate fractions. The CAS activity of the soluble fraction (cytosolic CAS activity) was 10 times higher than that of the particulate fraction. The CAS activity of the particulate fraction was confirmed to be localized in the mitochondria. Both enzymatic activities were clearly separated by non-denaturing PAGE. The enzyme with cytosolic CAS activity has been extensively purified and separated 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 showed 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 converting cyanide and cysteine into H2S and beta-cyanoalanine was confirmed by the detection of significant 14CN incorporation into beta-cyanoalanine. These results indicated that cytosolic CAS activity is due to cytosolic CS and suggested that the CAS activity of CS is likely to be involved in cyanide metabolism in plant tissues.
Maruyama
A
A
Biological Institute, Graduate School of Science, Tohoku University, Senda, Japan.
Ishizawa
K
K
Takagi
T
T
Esashi
Y
Y
D000409:Alanine / Q000031:analogs & derivatives / Q000096:biosynthesis; D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008190:Lyases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D010944:Plants / Q000201:enzymology*; D012639:Seeds / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid
D008190:Lyases; C019955:beta-cyanoalanine synthase; D000409:Alanine