Low S Y, Taylor P M, Ahmed A, Pogson C I, Rennie M J
Department of Anatomy and Physiology, University of Dundee, Scotland, U.K.
Biochem J. 1991 Aug 15;278 ( Pt 1)(Pt 1):105-11. doi: 10.1042/bj2780105.
We investigated the effects of glutamine and histidine analogues on glutamine transport processes in membrane vesicles prepared from rat liver (sinusoidal membrane) and skeletal muscle (sarcolemma). L-[14C]Glutamine is transported in these membranes predominantly by Systems N/Nm (liver and muscle respectively), and to a lesser extent by Systems A and L (e.g. about 60, 20 and 20% of total flux respectively via Systems N, A and L at 0.05 mM-glutamine in liver membrane vesicles). The glutamine anti-metabolites 6-diazo-5-oxo-L-norleucine and acivicin were relatively poor inhibitors of glutamine uptake into liver membrane vesicles (less than 25% inhibition at 20-fold excess) and appeared primarily to inhibit System A activity (i.e. N-methylaminoisobutyric acid-inhibitable glutamine uptake). In similar experiments azaserine (also a glutamine anti-metabolite) inhibited approx. 50% of glutamine uptake, apparently by inhibition of System A and also of System L (i.e. 2-amino-2-carboxybicyclo[2,2,1]heptane-inhibitable glutamine uptake). Glutamate gamma-hydroxamate, aspartate beta-hydroxamate, histidine and N'-methylhistidine were all strong inhibitors of glutamine uptake into liver membrane vesicles (greater than 65% inhibition at 20-fold excess), but neither homoglutamine nor N'-methylhistidine produced inhibition. L-Glutamate-gamma-hydroxamate was shown to be a competitive inhibitor of glutamine transport via System N (Ki approximately 0.6 mM). Glutamine uptake in sarcolemmal vesicles showed a similar general pattern of inhibition as in liver membrane vesicles. The results highlight limits on the substrate tolerance of System N; we suggest that the presence of both an L-alpha-amino acid group and a nitrogen group with a delocalized lone-pair of electrons (amide or pyrrole type), separated by a specific intramolecular distance (C2-C4 chain equivalent), is important for substrate recognition by this transporter.
我们研究了谷氨酰胺和组氨酸类似物对从大鼠肝脏（窦状膜）和骨骼肌（肌膜）制备的膜囊泡中谷氨酰胺转运过程的影响。L-[14C]谷氨酰胺在这些膜中主要通过系统N/Nm（分别为肝脏和肌肉）进行转运，在较小程度上通过系统A和L进行转运（例如，在肝膜囊泡中0.05 mM谷氨酰胺时，通过系统N、A和L的总通量分别约为60%、20%和20%）。谷氨酰胺抗代谢物6-重氮-5-氧代-L-正亮氨酸和阿西维辛对谷氨酰胺摄取到肝膜囊泡中的抑制作用相对较弱（20倍过量时抑制率小于25%），且主要表现为抑制系统A的活性（即N-甲基氨基异丁酸可抑制的谷氨酰胺摄取）。在类似实验中，重氮丝氨酸（也是一种谷氨酰胺抗代谢物）抑制了约50%的谷氨酰胺摄取，显然是通过抑制系统A以及系统L（即2-氨基-2-羧基双环[2,2,1]庚烷可抑制的谷氨酰胺摄取）。谷氨酸γ-羟肟酸、天冬氨酸β-羟肟酸、组氨酸和N'-甲基组氨酸都是谷氨酰胺摄取到肝膜囊泡中的强抑制剂（20倍过量时抑制率大于65%），但高谷氨酰胺和N'-甲基组氨酸均未产生抑制作用。L-谷氨酸γ-羟肟酸被证明是通过系统N转运谷氨酰胺的竞争性抑制剂（Ki约为0.6 mM）。肌膜囊泡中的谷氨酰胺摄取显示出与肝膜囊泡中类似的总体抑制模式。结果突出了系统N对底物耐受性的限制；我们认为，存在一个L-α-氨基酸基团和一个具有离域孤对电子的氮基团（酰胺或吡咯类型），并由特定的分子内距离（相当于C2-C4链）隔开，对于该转运体识别底物很重要。
