Wheeler T J
Department of Biochemistry, University of Louisville School of Medicine, KY 40292.
Biochim Biophys Acta. 1989 Mar 13;979(3):331-40. doi: 10.1016/0005-2736(89)90253-8.
Three compounds which inhibit glucose transport in rat adipocytes have been proposed to act directly on the glucose transporter protein. We tested these proposals by examining the effects of the compounds on the stereospecific glucose uptake catalyzed by adipocyte membrane proteins after reconstitution into liposomes. Effects on the transport activity reconstituted from human erythrocyte membranes were also examined. Glucose 6-phosphate, which was suggested to inhibit the transporter noncompetitively (Foley, J.E. and Huecksteadt, T.P. (1984) Biochim. Biophys. Acta 805, 313-316), had no effect on either type of reconstituted transporter, even when present at 5 mM on both sides of the liposomal membranes. Thus, it is unlikely to act directly on the transporter. The metalloendoproteinase substrate dipeptide Cbz-Gly-Phe-NH2, which inhibited insulin-stimulated but not basal glucose uptake in adipocytes (Aiello, L.P., Wessling-Resnick, M. and Pilch, P.F. (1986) Biochemistry 25, 3944-3950), inhibited the reconstituted erythrocyte transporter noncompetitively with a Ki of 1.5-2 mM. The inhibition of the erythrocyte transporter was identical in liposomes of soybean and egg lipid. Transport reconstituted using adipocyte membrane fractions was also inhibited by the dipeptide, with the activity from basal microsomes more sensitive than that from insulin-stimulated plasma membranes. These results indicate that the dipeptide interacts directly with the transporter, and may be a potentially useful probe for changes in transporter structure accompanying insulin action. Phenylarsine oxide, which was suggested to act directly on the adipocyte transporter (Douen, A.G., and Jones, M.N. (1988) Biochim. Biophys. Acta 968, 109-118), produced only slight (about 10%) inhibition of the reconstituted adipocyte and erythrocyte transporters, even when present at 100-200 microM and after 30 min of pretreatment. These results suggest that the major actions of phenylarsine oxide observed in adipocytes are not direct effects on the transporter, but rather effects on the pathways by which insulin regulates glucose transport activity (Frost, S.C. and Lane, M.D. (1985) J. Biol. Chem. 260, 2646-2652).
三种能抑制大鼠脂肪细胞葡萄糖转运的化合物被认为是直接作用于葡萄糖转运蛋白。我们通过检测这些化合物对重构到脂质体中的脂肪细胞膜蛋白催化的立体特异性葡萄糖摄取的影响，来验证这些推测。我们还检测了它们对由人红细胞膜重构的转运活性的影响。葡萄糖-6-磷酸被认为能非竞争性抑制转运蛋白（Foley, J.E.和Huecksteadt, T.P.（1984年）《生物化学与生物物理学报》805卷，313 - 316页），但即使在脂质体膜两侧都存在5 mM的情况下，它对两种重构的转运蛋白都没有影响。因此，它不太可能直接作用于转运蛋白。金属内蛋白酶底物二肽Cbz - Gly - Phe - NH₂能抑制脂肪细胞中胰岛素刺激的而非基础的葡萄糖摄取（Aiello, L.P., Wessling - Resnick, M.和Pilch, P.F.（1986年）《生物化学》25卷，3944 - 3950页），它对重构的红细胞转运蛋白有非竞争性抑制作用，抑制常数Ki为1.5 - 2 mM。在大豆脂质和鸡蛋脂质的脂质体中，对红细胞转运蛋白的抑制作用相同。使用脂肪细胞膜组分重构的转运也受到该二肽的抑制，基础微粒体的活性比胰岛素刺激的质膜的活性更敏感。这些结果表明该二肽直接与转运蛋白相互作用，可能是伴随胰岛素作用的转运蛋白结构变化的潜在有用探针。苯胂酸被认为直接作用于脂肪细胞转运蛋白（Douen, A.G.和Jones, M.N.（1988年）《生物化学与生物物理学报》968卷，109 - 118页），但即使在100 - 200 microM存在并预处理30分钟后，它对重构的脂肪细胞和红细胞转运蛋白也只有轻微（约10%）的抑制作用。这些结果表明在脂肪细胞中观察到的苯胂酸的主要作用不是对转运蛋白的直接影响，而是对胰岛素调节葡萄糖转运活性的途径的影响（Frost, S.C.和Lane, M.D.（1985年）《生物化学杂志》260卷，2646 - 2652页）。
