D. Left: Anti-FLAG precipitates from HEK293 cells transiently expressing FLAG-tagged CDKL5 (wild type "WT" or a K42R kinase-dead "KD" mutant) were incubated with the synthetic peptides indicated, in the presence of [γ-32P]-labelled ATP-Mg2+ and peptide phosphorylation was measured by Cerenkov counting. Data are represented as mean ± SEM from three independent experiments. Right: Same but anti-FLAG precipitates were subjected to SDS-PAGE and autoradiography to detect CDKL5 autophosphorylation, or western blotting with CDKL5-pTyr171 antibody specific for the CDKL5-Tyr171 autophosphorylation site 