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. 2016 Aug 5;283(18):3371–3388. doi: 10.1111/febs.13811

Figure 3.

Figure 3

Production of ENPP recombinant proteins. (A) Representative and schematic illustrations of recombinant mouse ENPP2‐1‐Target (mENPP2‐1‐T) chimera purified as described in Kato et al. [42,43] and recombinant 8xHistidine tag version (mENPP2‐1‐8xHis). (B) Flow chart for the purification of mENPP2‐1‐T and mENPP2‐1‐8xHis. (C) Left panel, the purity of recombinant mENPP2‐1‐T and mENPP2‐1‐8xHis enzymes was analysed using separation of 20 μm of protein on an SDS/PAGE gel followed by staining with Coomassie. Right panel, 20 μm of mENPP2‐1‐T was probed by anti‐6xHis western blot. (D) 1 μg of mENPP2‐1‐T and mENPP2‐1‐8xHis enzymes were used as substrates for PNGase F and Endo H deglycosylation enzymes. Samples were resolved on SDS/PAGE and stained by Coomassie. Black star indicates glycosylated ENPP1, green star indicates deglycosylated ENPP1, red star indicates deglycosylation resistant ENPP1.