Pathogenic variants of human Aromatic L-Amino Acid Decarboxylase: evidences of misfolding in functionally active variants

Dopa decarboxylase (DDC), a homodimeric pyridoxal 5’-phosphate (PLP)-dependent enzyme, is responsible for the decarboxylation of L-Dopa and L-5-hydroxytryptophan to neurotransmitters dopamine and serotonin, respectively. Missense mutations of the DDC gene lead to a rare and often fatal form of inborn infantile Parkinsonism, known as AADC deficiency 1. Up till now, it was shown that an incorrect apo→holo DDC transition is the reason of the pathogenicity of the majority of the mutations so far examined, with the exception of the R347 mutations defective in catalysis 2. Here we extend our work to a group of pathogenic mutations concerning residues distributed in different regions of DDC: P47H on the N-terminus, E283A, R285A, P210L and W267R on the large domain and R412W on the small C-terminal domain. The variants in their purified recombinant form exhibit kcat and Km values for L-Dopa as well as equilibrium binding dissociation constant values for PLP not very different from those of the wild-type. Thus, their functional properties do not seem to be responsible for their pathogenicity. Nonetheless, CD and DLS experiments highlight that all of these variants display even at different extent (i) an alteration of their tertiary structure and of their surface exposure features, and (ii) a propensity to an electrostaticdriven aggregation, mainly in their apo form. These data, together with the fact that these variants are expressed in E.coli at a level less than 1% of the wild-type, strongly suggest that folding defects characterize the variants. Thus, their inability to achieve or maintain a fully functional conformation would be the reason of their pathogenicity. Our results expand the knowledge of the enzymatic phenotypes leading to AADC deficiency.