Cloning and characterization of Clonorchis sinensis glucose transporter 2 (CsGLUT2)

초록

Clonorchis sinensis (Cs) infects the body when freshwater fish is ingested raw or undercooked, causing clonorchiasis, cholangitis, and digestive symptoms. C. sinensis utilizes exogenous glucose for its energy production. Although we have reported a glucose transporter, the NCBI database shows existing several isoforms of glucose transporters. Therefore, we try to isolate other one glucose transporter in Cs. A complementary DNA (cDNA) encoding a Cs glucose transporter 2 (CsGLUT2) was isolated from the adult Cs total RNA using overlap extension PCR method. The open reading frame of CsGLUT2 cDNA consists of 1491 base pairs that encode a 497-amino acid residue protein. Hydropathy analysis suggested that CsGLUT2 possess 12 putative membrane spanning domains. When expressed in Xenopus laevis oocytes, CsGLUT2 mediated the transport of radiolabeled deoxy-D-glucose in a time-dependent but sodium-independent manner. Concentration dependency results showed saturable kinetics and followed the Michaelis-Menten equation. Nonlinear regression analyses yielded a Km value of 2.7 mM and 588.5±53.0 μM and a Vmax value of 476 pmol/oocyte/ hr for [1,2-3H]2-deoxy-D glucose. No trans-uptakes of a-ketoglutarate, p-aminohippurate, taurocholate, argine, and carnitine were observed. There is no exchanger properties by efflux experiments. When inhibition experiment was performed using several kinds of hexose, only galactose showed weak inhibition. Considering obtained results, this glucose transport protein may contribute to the molecular biological study of carbohydrate metabolism and new drug development of C. sinensis. rogen metabolite movement in Cs.