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. 2025 Jul 6;34(17):1505–1516. doi: 10.1093/hmg/ddaf100

Figure 2.

Figure 2

acad8−/−;acadsb−/− fish have a discernable phenotype with increased metabolite accumulation. (A) Lateral view of acad8−/− and acadsb−/− fish at 6 months. No discernable phenotype in acad8−/− and acadsb−/− was noted. (B) C4-acylcarnitine and C5-acylcranitine measurements in liver at 6 month old adult zebrafish. In spite of no phenotypic differences, C5-acylcarnitine was elevated in acadsb−/−. (C) Lateral view of acad8 and acadsb double mutants at 29 dpf. No discernable phenotypes were found in both acad8+/−;acadsb−/− and acad8−/−;acadsb+/− but significant developmental delay in acad8−/−;acadsb−/− compound mutants was noted. Scale represents 1 mm in size. (D) Mortality of acad8 −/−;acadsb−/− compound mutant from two different incrosses; acad8+/−;acadsb−/− and acad8−/−;acadsb+/−. Lethality of acad8−/−;acadsb−/− from two different crosses begins equally at 29 dpf with significant loss at 45 dpf. (E) Elevation of C4-acylcarnitine levels in acad8−/−;acadsb−/− at 29 dpf. C4 acylcarnitine levels corresponding to the expected inability to process isobutyryl-CoA (the substrate for acad8) were significantly elevated in acad8−/−;acadsb−/− zebrafish, but very mildly increased in acad8−/−. (F) Elevation of C5-acylcarnitine levels in acadsb−/− and acad8 −/−;acadsb −/− at 29 dpf. C5 acylcarnitine levels corresponding to the expected inability to process 2-methylbutyryl-CoA (the substrate for acadsb) and were elevated in acadsb−/− and even higher in acad8−/−; acadsb−/− fish. (G, H) Predicted zebrafish binding pocket model. IBD can accommodate its native substrate, isobutyryl-CoA, but due to its relatively small size, it is unable to bind a bulkier molecule of 2-methylbutyryl-CoA. The circle denotes the overhang of the additional carbon present in 2-methylbutyryl-CoA outside of the binding pocket. Based on this predictive model, a larger binding pocket of zebrafish 2MBD is predicted to accommodate both butyryl-CoA (purple) and 2-methylbutyryl-CoA. ns, not significant; **(P < 0.005); ***(P < 0.001); ****(P < 0.0001). WT line in a, B, E, and F are normal TAB5 strain.