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. 2023 Sep 15;24(18):14154. doi: 10.3390/ijms241814154

Figure 3.

Figure 3

Expression of the putative bony fish ALOX15 orthologs in the baculovirus insect cell system. Putative ALOX15 orthologs of selected bony fish species were expressed as N-terminal His-tag fusion proteins in Sf9 cells and aliquots of the cell lysate supernatants were analyzed by Western-blot analysis using an anti-his-tag fusion protein antibody. (M) Molecular weight markers. (A) Quantification of the immune signals. Here, known amounts (lane 1, 500 ng; lane 2, 250 ng) of pure recombinant N-terminal his-tag fusion ALOX of Myxococcus fulvus were applied. (B) Expression of putative bony fish ALOX15 orthologs (lanes 3–5). Lane 3: putative ALOX15 ortholog of N. furzeri, 100 µg lysate supernatant protein; lane 4, putative ALOX15 ortholog of P. nyererei, 100 µg lysate supernatant protein; lane 5, putative ALOX15 ortholog of S. formosus, 100 µg lysate supernatant protein. Expressing the putative ALOX15 ortholog of S. formosus, we observed an additional immunoreactive band with a molecular weight of about 30 kDa (lane 5). The identity of this additional band remains unclear. (C) Sf9 cell lysate supernatant (100 µg total lysate supernatant protein) that was infected with the non-lipoxygenase control baculovirus (lane 6).