Figure 3. In vivo substrate affinity assay.
A) Schematic of the in vivo substrate affinity assay
B) Competitor substrate set of SPK-, SPQ- and PPK-GFP is efficiently expressed and N-terminally methylated after 32h of expression following calcium phosphate transfection.
C) The efficiency of N-terminal methylation of immunoprecipitated SPK- and PPK-GFP-FLAG test substrates is variable and depends on the presence of co-transfected competitor substrate.
D) Quantification of the efficiency of N-terminal methylation of SPK- and PPK-GFP-FLAG test substrates in the presence of SPK-, SPQ- or PPK-GFP competitor. Data were normalized and compared to the level of N-terminal methylation of test substrates co-expressed in the presence of unmethylatable SPQ-GFP competitor substrate by two-tailed independent t tests. * indicates P<0.01. n=3 independent repetitions per combination. Error bars: +/− 1 s.d.