Ebert Karolin, Ludwig Maren, Geillinger Kerstin Elisabeth, Schoberth Gina Catalina, Essenwanger Jasmin, Stolz Jürgen, Daniel Hannelore, Witt Heiko
Pediatric Nutritional Medicine, Klinikum rechts der Isar (MRI), Else Kröner-Fresenius-Zentrum für Ernährungsmedizin (EKFZ), Technische Universität München, Fachgebiet Pädiatrische Ernährungsmedizin, Gregor-Mendel-Str. 2, 85354, Freising, Germany.
ZIEL-Institute for Food and Health, Freising, Germany.
J Membr Biol. 2017 Apr;250(2):171-182. doi: 10.1007/s00232-016-9945-7. Epub 2017 Jan 12.
Although increased dietary fructose consumption is associated with metabolic impairments, the mechanisms and regulation of intestinal fructose absorption are poorly understood. GLUT5 is considered to be the main intestinal fructose transporter. Other GLUT family members, such as GLUT7 and GLUT9 are also expressed in the intestine and were shown to transport fructose and glucose. A conserved isoleucine-containing motif (NXI) was proposed to be essential for fructose transport capacity of GLUT7 and GLUT9 but also of GLUT2 and GLUT5. In assessing whether human GLUT2, GLUT5, GLUT7, and GLUT9 are indeed fructose transporters, we expressed these proteins in Xenopus laevis oocytes. Stably transfected NIH-3T3 fibroblasts were used as second expression system. In proving the role of the NXI motif, variants p.I322V of GLUT2 and p.I296V of GLUT5 were tested as well. Sugar transport was measured by radiotracer flux assays or by metabolomics analysis of cell extracts by GC-MS. Fructose and glucose uptakes by GLUT7 were not increased in both expression systems. In search for the physiological substrate of GLUT7, cells overexpressing the protein were exposed to various metabolite mixtures, but we failed to identify a substrate. Although urate transport by GLUT9 could be shown, neither fructose nor glucose transport was detectable. Fructose uptake was decreased by the GLUT2 p.I322V variant, but remained unaffected in the p.I296V GLUT5 variant. Thus, our work does not find evidence that GLUT7 or GLUT9 transport fructose or glucose or that the isoleucine residue determines fructose specificity. Rather, the physiological substrate of GLUT7 awaits to be discovered.
尽管饮食中果糖摄入量的增加与代谢障碍有关，但肠道果糖吸收的机制和调节仍知之甚少。GLUT5被认为是主要的肠道果糖转运体。其他GLUT家族成员，如GLUT7和GLUT9也在肠道中表达，并被证明可转运果糖和葡萄糖。一个保守的含异亮氨酸基序（NXI）被认为对GLUT7和GLUT9以及GLUT2和GLUT5的果糖转运能力至关重要。在评估人类GLUT2、GLUT5、GLUT7和GLUT9是否确实是果糖转运体时，我们在非洲爪蟾卵母细胞中表达了这些蛋白。稳定转染的NIH-3T3成纤维细胞被用作第二个表达系统。为了证明NXI基序的作用，还测试了GLUT2的p.I322V变体和GLUT5的p.I296V变体。通过放射性示踪剂通量测定或通过GC-MS对细胞提取物进行代谢组学分析来测量糖转运。在两个表达系统中，GLUT7对果糖和葡萄糖的摄取均未增加。为了寻找GLUT7的生理底物，将过表达该蛋白的细胞暴露于各种代谢物混合物中，但我们未能鉴定出底物。尽管可以显示GLUT9介导尿酸转运，但未检测到果糖或葡萄糖转运。GLUT2的p.I322V变体使果糖摄取减少，但p.I296V GLUT5变体中的果糖摄取未受影响。因此，我们的研究没有发现证据表明GLUT7或GLUT9转运果糖或葡萄糖，或者异亮氨酸残基决定果糖特异性。相反，GLUT7的生理底物有待发现。
