Expression and localization of recombinant human EDG-1 receptors in Pichia pastoris.
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.
Yang
Guangxiao
G
Research Centre of Gene Diversity and Designed Agriculture, Ministry of Education Key Laboratory for Biodiversity Science and Ecological Engineering, School of Life Science, Fudan University, Shanghai 200433, P.R. China.
Liu
Tianlei
T
Peng
Wulin
W
Sun
Xiaobo
X
Zhang
Han
H
Wu
Chaoqun
C
Shen
Daleng
D
D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence; D010843:Pichia / Q000235:genetics*; D011869:Radioligand Assay; D049349:Receptors, Lysosphingolipid / Q000096:biosynthesis* / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
D049349:Receptors, Lysosphingolipid; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins