Conditional Expression of AATK Variants

AATK coding sequence was kindly provided by Ma and Rubin [16 (link)] and cloned into, pEYFP-C2 (Clontech) and pCMVTag1 (Flag; Agilent). The ATP binding pocket of AATK was mutated (Ala>Lys) by site-directed mutagenesis (Agilent). DNA of AATK wt or AATK KD were cloned into pcDNATo/Myc/His vector (Thermo Fisher Scientific). TREx293 cells, that stably express the Tet repressor (Thermo Fisher Scientific), were transfected with the expression vector pcDNA4TO-AATK wt or pcDNA4TO-AATK KD and selected with Zeocin (Thermo Fisher Scientific). The cells were cultivated in DMEM with 10% tetracycline-free serum (Biochrom) and 1% penicillin and streptomycin (GIBCO). The selection of the clones was performed using blasticidin (5 µg/ml, Roth) and Zeocin (500 µg/ml, Thermo Fisher Scientific). The induction of AATK wt or AATK KD was performed using doxycycline (2 µg/ml, Thermo Fisher Scientific).