Figure 1

Sequence, domains and expression of GPKOW. A. Sequence alignment of GPKOW from human (NCBI NP_056513.2, upper line), monkey (NCBI XP_001105674, middle line) and cattle (NCBI XP_616889, lower line). The G-patch domain is indicated in yellow and the two KOW motifs are indicated in red. Two potential PKA phosphorylation sites, S27 and T316 (according to human GPKOW), are indicated in green. B. The G-patch domain of GPKOW as defined in the ELM database. Upper case letters illustrate the G-patch consensus sequence, where bold letters are glycines, underscored letters are hydrophobic residues and aromatic amino acids are marked in italic. C. 293T cells were left untreated (lanes 1, 3 and 5) or transfected with full-length GPKOW (lanes 2, 4 and 6) or GPKOW with a C-terminal V5-tag (lane 7). U2OS cells were left untreated (lane 8) or transfected with full-length GPKOW (lane 9). Twenty hours post transfection the cells were harvested and lysed. Eighteen μg (lanes 1-4 and 7), 36 μg (lane 5-6) or 46 μg (lanes 8-9) of protein was loaded per lane, separated by SDS-PAGE (10% gels) and analyzed by immunoblotting using three different GPKOW antibodies, 1894 (lanes 1-2 and 8-9), 0287 (lanes 3-4) and B01 (lanes 5-6) or V5 antibody (lane 7). One representative immunoblot from each experiment is presented.