Cloning and expression of human placental L-Dopa decarboxylase

L-Dopa decarboxylase (DDC) is a pyridoxal 5-phosphate (PLP)-dependent enzyme that catalyses the decarboxylation of L-Dopa to dopamine. In this study we show the expression of DDC in human placental tissue and present data on the molecular cloning and in vitro expression of the active recombinant enzyme. Our analyses indicated the presence of both alternative DDC mRNA splice variants (neuronal and nonneuronal) in human placenta. Cloning of the coding region of the DDC cDNA into the pTrcHisA expression vector led to the production of the enzymatically active recombinant protein. The obtained recombinant enzyme specific activity values were in good agreement with the results obtained for the purified enzyme from human kidney. The availability of active recombinant human DDC could provide information leading to the better understanding of the enzyme's structure and substrate specificity, as well as its regulation and involvement in pathological conditions.     

<Emphasis Type="SmallCaps">l</Emphasis>-Dopa decarboxylase expression profile in human cancer cells

l-Dopa decarboxylase (DDC) catalyses the decarboxylation of l-Dopa. It has been shown that the DDC gene undergoes alternative splicing within its 5′-untranslated region (UTR), in a tissue-specific manner, generating identical protein products. The employment of two alternative 5′UTRs is thought to be responsible for tissue-specific expression of the human DDC mRNA. In this study, we focused on the investigation of the nature of the mRNA expression in human cell lines of neural and non-neural origin. Our results show the expression of a neural-type DDC mRNA splice variant, lacking exon 3 in all cell lines studied. Co-expression of the full length non-neural DDC mRNA and the neural-type DDC splice variant lacking exon 3 was detected in all cell lines. The alternative DDC protein isoform, Alt-DDC, was detected in SH-SY5Y and HeLa cells. Our findings suggest that the human DDC gene undergoes complex processing, leading to the formation of multiple mRNA isoforms. The study of the significance of this phenomenon of multiple DDC mRNA isoforms could provide us with new information leading to the elucidation of the complex biological pathways that the human enzyme is involved in.