RAB12 and functionally investigated variants. (a) Schematic view of the RAB12 protein with the identified mutations p.Gly13Asp and p.Ile196Val in MD patients (red arrows), the guanosine diphosphate/guanosine triphosphate (GDP/GTP)-binding sites (red), the predicted effector region (blue), and the posttranslational modifications on amino acid positions 1 (N-acetylation), 21, 25, 106 (phosphorylations), 243, and 244 (geranylgeranylations) are indicated. RefSeq: NM_001025300.2, NP_001020471; (b) The Glycine at position 13 (left panel, red) and the Isoleucine at position 196 (right panel, red) are conserved across different species; (c) Proteins of transfected SH-SY5Y cells were incubated with GTP for 1 h and PO43− production was measured. GTPase activity of mutant RAB12 expressing cells was normalized for GTPase activity of wild-type (WT) RAB12 expressing cells. The bars indicate the means ± standard error of mean, One-way analysis of variance (ANOVA) with Bonferroni’s post-hoc test. ** p < 0.01, n = 4; (d) Homology model of active/GTP-bound RAB12 in ribbon representation. WT is presented in black (left model) and p.Ile196Val in blue (right model). Magnesium ions are shown as green spheres, amino acid residue 196 is marked by red points, and GTP is indicated by sticks. The figures were generated with PyMOL (PyMOL Molecular Graphics System, Version 1.7.0 Schrödinger, LLC, New York, NY, USA).