Machín Felix, Medina Braulio, Navarro Francisco J, Pérez M Dolores, Veenhuis Marten, Tejera Paula, Lorenzo Helena, Lancha Ana, Siverio José M
Departamento de Bioquímica y Biología Molecular, Grupo del Metabolismo del Nitrógeno, Universidad de La Laguna, Tenerife, Canarias, Spain.
Yeast. 2004 Feb;21(3):265-76. doi: 10.1002/yea.1075.
Ynt1 is the only high-affinity nitrate uptake system in Hansenula polymorpha. Nitrate uptake was directly correlated with the Ynt1 levels and shown to be independent of nitrate reductase (NR) activity levels. Ynt1 failed to transport chlorate and, as a result, strains lacking YNT1 were sensitive to chlorate, as is the wild-type. Nitrite uptake in a wild-type strain was partially inhibited by nitrate to levels shown by a YNT1-disrupted strain in which, in turn, nitrite transport was not inhibited by nitrate. It is concluded that nitrite uptake takes place by two different transport systems: Ynt1 and a nitrite-specific transporter(s). The nitrite-specific transport system was induced by nitrate; consistently, no induction was observed in strains lacking the transcription factor YNA1, which is involved in nitrate and nitrite induction of the nitrate assimilatory structural genes. Ynt1 presents its optimal rate for nitrite uptake at pH 6, while pH 4 was optimal for the specific nitrite uptake system(s). At pH 5.5, the contribution of Ynt1 to high-affinity nitrate and nitrite uptake was around 95% and 60%, respectively. The apparent Km of Ynt1 for nitrate and nitrite is in the microM range, as is the specific nitrite uptake system for nitrite. The analysis of the effect of the reduced nitrogen sources on nitrate assimilation revealed that glutamine inactivates nitrate and nitrite transport, dependent on Ynt1, but not the nitrite-specific system.
Ynt1是多形汉逊酵母中唯一的高亲和力硝酸盐摄取系统。硝酸盐摄取与Ynt1水平直接相关，且显示与硝酸还原酶（NR）活性水平无关。Ynt1无法转运氯酸盐，因此，缺乏YNT1的菌株对氯酸盐敏感，野生型菌株也是如此。野生型菌株中的亚硝酸盐摄取受到硝酸盐的部分抑制，抑制程度与YNT1缺失菌株相同，而在YNT1缺失菌株中，亚硝酸盐转运不受硝酸盐抑制。由此得出结论，亚硝酸盐摄取通过两种不同的转运系统进行：Ynt1和一种亚硝酸盐特异性转运体。亚硝酸盐特异性转运系统由硝酸盐诱导；一致的是，在缺乏转录因子YNA1的菌株中未观察到诱导现象，YNA1参与硝酸盐同化结构基因的硝酸盐和亚硝酸盐诱导。Ynt1在pH 6时亚硝酸盐摄取速率最佳，而pH 4是特异性亚硝酸盐摄取系统的最佳pH值。在pH 5.5时，Ynt1对高亲和力硝酸盐和亚硝酸盐摄取的贡献分别约为95%和60%。Ynt1对硝酸盐和亚硝酸盐的表观Km值在微摩尔范围内，特异性亚硝酸盐摄取系统对亚硝酸盐的表观Km值也是如此。对还原氮源对硝酸盐同化影响的分析表明，谷氨酰胺会使依赖Ynt1的硝酸盐和亚硝酸盐转运失活，但不会使亚硝酸盐特异性系统失活。
