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Characterization of Recombinant Human Aromatic l ‐Amino Acid Decarboxylase Expressed in COS Cells
Author(s) -
Sumi Chiho,
Ichinose Hiroshi,
Nagatsu Toshiharu
Publication year - 1990
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 - 10.1111/j.1471-4159.1990.tb04601.x
Subject(s) - decarboxylation , aromatic l amino acid decarboxylase , complementary dna , enzyme , recombinant dna , biochemistry , substrate (aquarium) , transfection , microbiology and biotechnology , chemistry , amino acid , carboxy lyases , pyridoxal , pyridoxal phosphate , stereochemistry , biology , catalysis , gene , cofactor , ecology
The expression vector containing the full‐length cDNA of human aromatic l ‐Amino acid decarboxylase (EC 4.1.1.28) was transfected in COS cells by a modified calcium phosphate coprecip‐itation method. The cells transfected with plasmids that had a true direction of the cDNA gave a major immunoreactive band at 50 kDa. This expressed enzyme catalyzed the decarboxylation of l‐3,4‐dihydroxyphenylalanine (l‐DOPA). l‐5‐hydroxytryptophan (l‐5‐HTP) and l‐ threo ‐3,4‐dihydroxyphenylserine. The optimal pH of the enzyme activity with l‐DOPA as a substrate was 6.5, whereas the enzyme had a broad pH optimum when l‐5‐HTP was used as a substrate. Addition of pyridoxal phosphate to the incubation mixture greatly enhanced the activity for both l‐DOPA and l‐5‐HTP.