Cystathionine gamma-lyase of Saccharomyces cerevisiae: structural gene and cystathionine gamma-synthase activity.
Purification of Saccharomyces cerevisiae cystathionine gamma-lyase (gamma-CTLase) was hampered by the presence of a protein migrating very close to it in various types of column chromatography. The enzyme and the contaminant were nevertheless separated by polyacrylamide gel electrophoresis. N-terminal amino acid sequence analysis indicated that they are coded for by CYS3 (CYI1) and MET17 (MET25), respectively, leading to the conclusion that CYS3 is the structural gene for gamma-CTLase and that the contaminant is O-acetylserine/O-acetylhomoserine sulfhydrylase (OAS/OAH SHLase). Based on these findings, we purified gamma-CTLase by the following strategy: (1) extraction of OAS/OAH SHLase from a CYS3-disrupted strain; (2) preparation of antiserum against it; (3) identification of a strain devoid of the OAS/OAH SHLase protein using this antiserum; and (4) extraction of gamma-CTLase from this strain. Purified gamma-CTLase had cystathionine gamma-synthase (gamma-CTSase) activity if O-succinylhomoserine, but not O-acetylhomoserine, was used as substrate. From this notion we discuss the evolutional relationship between S. cerevisiae gamma-CTLase and Escherichia coli gamma-CTSase.
Ono
B
B
Laboratory of Environmental Hygiene Chemistry, Faculty of Pharmaceutical Sciences, Okayama University, Japan.
Ishii
N
N
Naito
K
K
Miyoshi
S
S
Shinoda
S
S
Yamamoto
S
S
Ohmori
S
S
D000595:Amino Acid Sequence; D019757:Carbon-Oxygen Lyases; D002846:Chromatography, Affinity; D003542:Cystathionine gamma-Lyase / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D005800:Genes, Fungal / Q000235:genetics*; D007700:Kinetics; D008190:Lyases / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
C022696:O-succinylhomoserine (thiol)-lyase; D008190:Lyases; D019757:Carbon-Oxygen Lyases; D003542:Cystathionine gamma-Lyase