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Brain succinic semialdehyde dehydrogenase: identification of reactive lysyl residues labeled with pyridoxal‐5′‐phosphate
Author(s) -
Choi Soo Young,
Bahn Jae Hoon,
Lee Byung Ryong,
Jeon Seong Gyu,
Jang Joong Sik,
Kim Choong Kwon,
Jin Li Hua,
Kim Kyung Hee,
Park Jin Sun,
Park Jinseu,
Cho SungWoo
Publication year - 2001
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2001.00147.x
Subject(s) - pyridoxal , cofactor , biochemistry , chemistry , enzyme , pyridoxal phosphate , dehydrogenase , nad+ kinase , tetramer , stereochemistry
An NAD + ‐dependent succinic semialdehyde dehydrogenase from bovine brain was inactivated by pyridoxal‐5′‐ phosphate. Spectral evidence is presented to indicate that the inactivation proceeds through formation of a Schiff's base with amino groups of the enzyme. After NaBH 4 reduction of the pyridoxal‐5′‐phosphate inactivated enzyme, it was observed that 3.8 mol phosphopyridoxyl residues were incorporated/enzyme tetramer. The coenzyme, NAD + , protected the enzyme against inactivation by pyridoxal‐5′‐phosphate. The absorption spectrum of the reduced and dialyzed pyridoxal‐5′‐phosphate‐inactivated enzyme showed a characteristic peak at 325 nm, which was absent in the spectrum of the native enzyme. The fluorescence spectrum of the pyridoxyl enzyme differs completely from that of the native enzyme. After tryptic digestion of the enzyme modified with pyridoxal‐5′‐phosphate followed by [ 3 H]NaBH 4 reduction, a radioactive peptide absorbing at 210 nm was isolated by reverse‐phase HPLC. The sequences of the peptide containing the phosphopyridoxyllysine were clearly identical to sequences of other mammalian succinic semialdehyde dehydrogenase brain species including human. It is suggested that the catalytic function of succinic semialdehyde dehydrogenase is modulated by binding of pyridoxal‐5′‐phosphate to specific Lys 347 residue at or near the coenzyme‐binding site of the protein.