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. 2014 Feb 23;71(18):3611–3626. doi: 10.1007/s00018-014-1591-6

Fig. 2.

Fig. 2

DPP9-l is active. a Table summarizing the calculated K cat, K m, and K cat /K m results for the cleavage of several DPP9 substrates by 12.5 nM recombinant DPP9-S and DPP9-l. Assays were performed in triplicates and repeated at least three times. Values were calculated using non-linear regression (GraphPad Inc. Prism software). b Elution profiles of 200 nM recombinant DPP9-l and DPP9-S on a gel exclusion chromatography, using analytical Superdex S200. As size standard, Bio-Rad’s gel filtration standard was used. Profiles were generated using GraphPad Inc. Prism software with ml on the x-axis and UV absorbance (mAU 280 nm) on the y-axis. This experiment was repeated three times, shown is a representative. c Scheme of HA-tagged DPP9-l and DPP9-S constructs used for overexpression in HeLa cells for subsequent immunofluorescences, subcellular fractionation experiments, and cleavage-activity assays. d DPP activity was measured from HeLa cell extracts (5 μg) expressing HA-tagged DPP9-l or DPP9-S. The experiment was performed in triplicates, including error bars. e Western blots of lysates (10 μg) from HeLa cells overexpressing HA-tagged DPP9-S or DPP9-l developed with DPP9 (total) antibodies to detect both isoforms, or DPP9 (long) antibodies to detect only DPP9-l. Tubulin was used as loading control. Shown are the blots for the corresponding activity assays shown in d