Molecular cloning and functional characterization of a glucose transporter (CsGLUT) in Clonorchis sinensis

초록

A cDNA encoding a glucose transporter of Clonorchis sinensis (CsGLUT) was isolated from adult C. sinensis cDNA library. The open reading frame of CsGLUT cDNA consists of 1653 base pairs that encode a 550-amino acid residue protein. Hydropathy analysis suggested that CsGLUT possess 12 putative membranespanning domains. The Northern blot analysis result using poly(A)+RNA showed a strong band at ~2.1 kb for CsGLUT. When expressed in Xenopus oocytes, CsGLUT mediated the transport of radiolabeled deoxy-D-glucose in a time-dependent but sodium-independent manner. Concentration-dependency results showed saturable kinetics and followed Michaelis-Menten equation. Nonlinear regression analyses yielded a Km value of 588.5±53.0 μM and a Vmax value of 1500.0±67.5 pmol/oocyte/30 min for [1,2-3H]2-deoxy-Dglucose. No trans-uptakes of bile acid (taurocholic acid), amino acids (tryptophan and arginine), or p-aminohippuric acid were observed. CsGLUT-mediated transport of deoxyglucose was significantly and concentration-dependently inhibited by radio unlabeled deoxyglucose and D-glucose. 3-OMethylglucose at 10 μM and 100 μM inhibited deoxyglucose uptake by ~50 % without concentration dependence. No inhibitory effects by galactose, mannose, and fructose were observed. This work may contribute to the molecular biological study of carbohydrate metabolism and new drug development of Clonorchis sinensis.