Contribution to Substrate Recognition of Two Aromatic Amino Acid Residues in Putative Transmembrane Segment 10 of the Yeast Sugar Transporters Gal2 and Hxt2*
Contribution to Substrate Recognition of Two Aromatic Amino Acid Residues in Putative Transmembrane Segment 10 of the Yeast Sugar Transporters Gal2 and Hxt2*
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DOI：
10.1074/jbc.273.44.29106
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发表时间：
1998-10
期刊：
影响因子：
--
通讯作者：
M. Kasahara;M. Maeda
中科院分区：
文献类型：
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作者：
M. Kasahara;M. Maeda
对酿酒酵母Gal 2半乳糖转运蛋白和Hxt 2葡萄糖转运蛋白嵌合体的研究表明，Tyr 446是Gal 2识别半乳糖所必需的，Trp 455是Gal 2识别半乳糖所必需的。与这一发现一致，分别用Tyr和Trp替换Hxt 2的相应Phe 431和Tyr 440残基允许Hxt 2转运半乳糖，表明推定的跨膜区段10中的两个氨基酸残基在半乳糖识别中起确定的作用（Kasahara，M.，Shimoda，E.，和Maeda，M.（1997）J.Biol.Chem.272，16721-16724）。用其它19种氨基酸中的任一种替换Gal 2的Trp 455显示将半乳糖转运活性降低至野生型Gal 2的半乳糖转运活性的0至<20%。类似地研究了Phe 431在Hxt 2中的作用。除了Phe之外，仅在位置431处的Tyr能够以<20%的降低水平支持葡萄糖转运活性。与此相反，用其他氨基酸替换Hxt 2的Tyr 440显示，除了Pro和带电氨基酸之外，大多数替换都支持葡萄糖转运活性。残基431在糖识别中的重要性在其中Tyr 440被Trp替换的修饰的Hxt 2中更明显。葡萄糖转运仅由431位的芳香族氨基酸Phe、Tyr和Trp支持，半乳糖转运仅由Tyr支持。这些结果表明，位于跨膜段10中间的芳香族氨基酸（Tyr 446在Gal 2和Phe 431在Hxt 2）在底物识别中起着关键作用的酵母糖转运蛋白家族Gal 2和Hxt 2属于。
The comprehensive study of chimeras between the Gal2 galactose transporter and the Hxt2 glucose transporter ofSaccharomyces cerevisiae has shown that Tyr446is essential and Trp455 is important for galactose recognition by Gal2. Consistent with this finding, replacement of the corresponding Phe431 and Tyr440 residues of Hxt2 with Tyr and Trp, respectively, allowed Hxt2 to transport galactose, suggesting that the two amino acid residues in putative transmembrane segment 10 play a definite role in galactose recognition (Kasahara, M., Shimoda, E., and Maeda, M. (1997) J. Biol. Chem. 272, 16721–16724). Replacement of Trp455 of Gal2 with any of the other 19 amino acids was shown to reduce galactose transport activity to between 0 and <20% of that of wild-type Gal2. The role of Phe431 in Hxt2 was similarly studied. Other than Phe, only Tyr at position 431 was able to support glucose transport activity, at the reduced level of <20%. In contrast, replacement of Tyr440 of Hxt2 with other amino acids revealed that most replacements, with the exception of Pro and charged amino acids, supported glucose transport activity. The importance of residue 431 in sugar recognition was more pronounced in a modified Hxt2 in which Tyr440 was replaced with Trp. Glucose transport was supported only by the aromatic amino acids Phe, Tyr, and Trp at position 431, and galactose transport was supported only by Tyr. These results suggest that an aromatic amino acid located in the middle of transmembrane segment 10 (Tyr446 in Gal2 and Phe431 in Hxt2) plays a critical role in substrate recognition in the yeast sugar transporter family to which Gal2 and Hxt2 belong.