Severi Emmanuele, Javelle Arnaud, Merrick Mike
Department of Biology, University of York, York, UK.
Mol Membr Biol. 2007 Mar-Apr;24(2):161-71. doi: 10.1080/09687860601129420.
The ammonium transport (Amt) proteins are a highly conserved family of integral membrane proteins found in eubacteria, archaea, fungi and plants. Genetic, biochemical and bioinformatic analyses suggest that they have a common tertiary structure comprising eleven trans-membrane helices with an N-out, C-in topology. The cytoplasmic C-terminus is variable in length but includes a core region of some 22 residues with considerable sequence conservation. Previous studies have indicated that this C-terminus is not absolutely required for Amt activity but that mutations that alter C-terminal residues can have very marked effects. Using the Escherichia coli AmtB protein as a model system for Amt proteins, we have carried out an extensive site-directed mutagenesis study to investigate the possible role of this region of the protein. Our data indicate that nearly all mutations fall into two phenotypic classes that are best explained in terms of two distinct effects of the C-terminal region on AmtB activity. Residues within the C-terminus play a significant role in normal AmtB function and the C-terminal region might also mediate co-operativity between the three subunits of AmtB.
铵转运（Amt）蛋白是一类高度保守的整合膜蛋白家族，存在于真细菌、古细菌、真菌和植物中。遗传学、生物化学和生物信息学分析表明，它们具有共同的三级结构，由11个跨膜螺旋组成，拓扑结构为N端向外、C端向内。细胞质C端的长度可变，但包括一个约22个残基的核心区域，序列保守性相当高。先前的研究表明，Amt活性并非绝对需要这个C端，但改变C端残基的突变可能会产生非常显著的影响。我们以大肠杆菌AmtB蛋白作为Amt蛋白的模型系统，进行了广泛的定点诱变研究，以探讨该蛋白这一区域可能发挥的作用。我们的数据表明，几乎所有的突变都分为两个表型类别，根据C端区域对AmtB活性的两种不同影响来解释最为恰当。C端内的残基在正常的AmtB功能中发挥重要作用，并且C端区域可能还介导了AmtB三个亚基之间的协同作用。
