Sumi C, Ichinose H, Nagatsu T
Department of Biochemistry, Nagoya University School of Medicine, Japan.
J Neurochem. 1990 Sep;55(3):1075-8. doi: 10.1111/j.1471-4159.1990.tb04601.x.
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 coprecipitation 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.
通过改良的磷酸钙共沉淀法，将含有人类芳香族L-氨基酸脱羧酶（EC 4.1.1.28）全长cDNA的表达载体转染至COS细胞中。用具有cDNA正确方向的质粒转染的细胞在50 kDa处出现一条主要的免疫反应带。这种表达的酶催化L-3,4-二羟基苯丙氨酸（L-DOPA）、L-5-羟色氨酸（L-5-HTP）和L-苏式-3,4-二羟基苯丝氨酸的脱羧反应。以L-DOPA为底物时，酶活性的最适pH为6.5，而以L-5-HTP为底物时，该酶具有较宽的最适pH范围。向孵育混合物中添加磷酸吡哆醛可显著增强对L-DOPA和L-5-HTP的活性。
