Expression and localization of recombinant human EDG-1 receptors in Pichia pastoris.

ABSTRACT

The receptor for human endothelial differentiation gene-1 protein (EDG-1) was C-terminally tagged with green fluorescent protein and expressed in the methylotrophic yeast, Pichia pastoris. EDG-1 expression was driven by the highly inducible alcohol oxidase 1 promoter. Expression of EDG-1 recombinant protein was detected by Western blot analysis and confocal microscopy. The recombinant EDG-1 receptor protein was located in the plasma membrane. Radioligand binding assays demonstrated that the EDG-1 receptors expressed in Pichia pastoris have specific and saturation binding of (32)P-labeled sphingosine 1-phosphate.