Purification and characterization of a novel cysteine synthase isozyme from spinach hydrated seeds.
A novel type of cysteine synthase (CSase, EC 4.2.99.8) isozyme, designated as CSase 1', was purified to homogeneity from hydrated spinach seeds. The enzyme had a molecular weight of 68,000 and consisted of two identical subunits of M(r), 34,000. The apparent K(m) for O-acetyl-L-serine was 8.33 mM and that for sulfide was 0.66 mM. The activity of CSase 1' was maintained when it was treated at 60 degrees C for 1 min. This novel enzyme was similar to CSases 1, 2, and 3 already purified from spinach leaves, in results of double immunodiffusion, molecular weight, subunit composition, K(m) values for O-acetyl-L-serine and sulfide, and heat stability. On the other hand, N-terminal amino acid sequence, effects of immunotitration, pH optimum, and effects of hydroxylamine on purified CSase 1' were different from those of the other CSases. Furthermore, it was found that CSases 2S and 3S isolated from hydrated spinach seeds were identical with the CSases 2 and 3 reported previously. It was also disclosed that CSases 1, 2, and 3 were localized in chloroplasts, cytosol, and mitochondria, respectively.
Yamaguchi
T
T
Department of Bioresources Chemistry, Faculty of Horticulture, Chiba University, Japan.
Zhu
X
X
Masada
M
M
D000595:Amino Acid Sequence; D003547:Cysteine Synthase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D019811:Hydroxylamine / Q000494:pharmacology; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018515:Plant Leaves / Q000201:enzymology; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D012639:Seeds / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D012995:Solubility; D018724:Spinacia oleracea / Q000201:enzymology*; D014867:Water / Q000737:chemistry
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D010940:Plant Proteins; D014867:Water; D019811:Hydroxylamine; D003547:Cysteine Synthase