Purification and characterisation of tyrosine decar☐ylase and aromatic- l-amino-acid decar☐ylase

Abstract

Microbial tyrosine decar☐ylase (EC 4.1.1.25) and mammalian aromatic- l-amino-acid decar☐ylase (EC 4.1.1.28) catalyse the formation of tyramine from l-tyrosine. These enzymes were characterised after isolation to purity by methods including fast polymer liquid chromatography (FPLC). Tyrosine decar☐ylase was isolated from Streptococcus faecalis by FPLC anion exchange chromatography (11-times purification; 72% recovery; 23.2 U/mg protein). FPLC on Phenyl-Superose resulted in purification to 115 U/mg protein. Aromatic- l-amino-acid decar☐ylase was isolated from pig kidney by ammonium sulfate fractionation. DEAE chromatography, and FPLC anion exchange chromatography (21-times purification; 22% recovery; 0.71 U/mg protein). By FPLC chromatofocusing, tyrosine decar☐ylase eluted at pH 4.3 and aromatic- l-amino-acid decar☐ylase at pH 5.0. Isoelectric focusing of tyrosine decar☐ylase gave two bands (p I 4.4 and 4.5). With pyridoxal 5′-phosphate removed by ultrafiltration, only one band (p I 4.4) appeared, and SDS polyacrylamide electrophoresis confirmed the purity. FPLC gel filtration resulted in molecular weights 143 000 and 86 000, respectively, for tyrosine decar☐ylase and aromatic- l-amino-acid decar☐ylase. In SDS electrophoresis, tyrosine decar☐ylase had the monomer molecular weight 75 000, showing a dimer structure for the enzyme.