Mitchell A M, Manley S W, Mortimer R H
Conjoint Endocrine Laboratory, Royal Brisbane Hospital, Queensland, Australia.
Mol Cell Endocrinol. 1994 May;101(1-2):203-10. doi: 10.1016/0303-7207(94)90236-4.
We studied the effect of a number of amino acids on uptake of L-triiodothyronine (T3) in the human choriocarcinoma cell line, JAR. Tryptophan inhibited saturable T3 uptake by about 57% without any significant effect on the non-saturable uptake. Michaelis constant (Km) for T3 uptake was 1.06 +/- 0.15 microM (n = 15) with the corresponding maximum velocity (Vmax) of 24.2 +/- 3.1 pmol/min/mg cellular protein. For tryptophan uptake the Km was 1.31 +/- 0.26 microM (n = 7) and Vmax was 166.4 +/- 35.7 pmol/min/mg protein. The kinetic parameters for both uptake processes were similar to those reported in normal placenta. Uptake of T3 was inhibited by tryptophan but not phenylalanine, but tryptophan uptake was inhibited both by T3 and phenylalanine. Inhibition of T3 uptake by tryptophan was dose dependent, with an inhibition constant (Ki) of 2.9 +/- 0.5 mM. Similarly, tryptophan uptake was inhibited by T3 and phenylalanine in a dose dependent way with Ki values of 4.9 +/- 0.5 microM and 15.6 +/- 4.8 microM respectively. Km for T3 uptake was significantly increased to 1.86 +/- 0.42 microM (n = 4) in the presence of 3 mM unlabelled tryptophan and, similarly, Km for tryptophan uptake was significantly increased to 9.91 +/- 2.57 microM (n = 3) in the presence of 5 microM unlabelled T3. Efflux of T3 was progressively inhibited by increasing concentrations of both ligands, i.e. was saturable. We conclude that there is mutual competitive inhibition between uptake systems for T3 and tryptophan in JAR cells, but the kinetic parameters of cross-inhibition of uptake by the substrates suggest that the carriers are distinct. T3 may be transported in JAR cells by at least two transport systems with differing substrate specificities. We also demonstrated the presence of a saturable membrane carrier mediating the efflux of T3 from the cells which was subject to trans-inhibition by T3 and tryptophan.
我们研究了多种氨基酸对人绒毛膜癌细胞系JAR摄取L-三碘甲状腺原氨酸（T3）的影响。色氨酸可使T3的饱和摄取抑制约57%，而对非饱和摄取无显著影响。T3摄取的米氏常数（Km）为1.06±0.15微摩尔（n = 15），相应的最大速度（Vmax）为24.2±3.1皮摩尔/分钟/毫克细胞蛋白。色氨酸摄取的Km为1.31±0.26微摩尔（n = 7），Vmax为166.4±35.7皮摩尔/分钟/毫克蛋白。两种摄取过程的动力学参数与正常胎盘报道的相似。T3的摄取受到色氨酸而非苯丙氨酸的抑制，但色氨酸的摄取受到T3和苯丙氨酸两者的抑制。色氨酸对T3摄取的抑制呈剂量依赖性，抑制常数（Ki）为2.9±0.5毫摩尔。同样，T3和苯丙氨酸以剂量依赖性方式抑制色氨酸摄取，Ki值分别为4.9±0.5微摩尔和15.6±4.8微摩尔。在存在3毫摩尔未标记色氨酸的情况下，T3摄取的Km显著增加至1.86±0.42微摩尔（n = 4），同样，在存在5微摩尔未标记T3的情况下，色氨酸摄取的Km显著增加至9.91±2.57微摩尔（n = 3）。两种配体浓度增加时，T3的流出均逐渐受到抑制，即具有饱和性。我们得出结论，JAR细胞中T3和色氨酸的摄取系统之间存在相互竞争性抑制，但底物对摄取的交叉抑制动力学参数表明载体是不同的。T3在JAR细胞中可能通过至少两种具有不同底物特异性的转运系统进行转运。我们还证明了存在一种可饱和的膜载体介导T3从细胞中流出，该载体受到T3和色氨酸的反抑制。
