(571an) Efficient Expression and Purification of Human Membrane Protein in Fat Body of Silkworm Larvae Using Bmnpv Bacmid and Characterization of Its Biological Property

Membrane proteins play important roles in biological system, and are targeted in the screening new medicines. We tried the production of membrane protein in the fat body of silkworm using BmNPV bacmid1). Full length human (pro)renin receptor (hPRR-wTM) and only extracellular domain of human (pro)renin receptor (hPRR-w/oTM) lacking the transmembrane and cytoplasmic domain were used as membrane protein in this work, and was successfully expressed in the fat body of silkworm larvae. To investigate the intrinsic structural and functional property of (pro)renin receptor, this membrane protein was solubilized from the membrane with dodecyl-â-D-maltoside, and get the protein with yield of more than 60 %. And both purified full-length hPRR and its extracellular domain were comparatively characterized in binding assays by real-time monitored surface plasmon resonance (SPR) assay with immobilized human renin on the cuvette. Our results demonstrate that the full-length hPRR showed the biological function of (pro)renin receptor judging by SPR assay. On the other hand, hPRR-w/oTM didn't show any binding property to human renin. This indicates that transmembrane domain of hPRR is indispensable for biological function of human (pro)renin receptor. And an unidentified factor in the fat body was presumed which may be plays an important role in the mechanism of binding of hPRR with human renin. This study is the first findings describing the interaction of full-length hPRR and its extracellular domain with human renin and may provide an important contribution to the study of hPRR.

1) Motohashi et al, Biochem Biophys Res Commun. 326 (2005) 564-569.