MOLECULAR CLONING & REGULATION OF HUMAN INDUCIBLE NITRIC OXIDE SYNTHASE GENE

我们感兴趣的是寻找氨基酸和结构 决定水通道蛋白(AQP)透水性的特征 蛋白质家族和更大的膜固有家族 蛋白质(MIP)。我们已经在酵母中表达了AQP1蛋白 分泌囊泡，也有重组酵母表达的AQP1 变成脂泡。我们已经证明了这种蛋白质起到了水的作用 在两个环境中都有通道。我们正在表达的过程中 AQP2和AQP3在同一酵母系统中。我们已经紧密地表达了两个 MIP家族的相关蛋白(FPS和GlpF)，尽管 在序列上同源，不要充当水的通道。我们想要 要找到特定的氨基酸或蛋白质结构域，它们在 水通道和非水通道，这可能对 测定水的渗透性。为此，我们计划分析 所有MIP和AQP蛋白的序列和二级结构 并寻找保守或可变的区域。以下网站： 然后可以评估潜在确定的水的渗透性 试验性地使用我们的酵母表达系统。序列比对 使用CGC软件可以进行二级结构预测 该程序包可从匹兹堡超级计算中心获得。

We are interested in finding the amino acids and structural features which determine water permeability in the aquaporin (AQP) family of proteins and in the larger family of membrane intrinsic proteins (MIPs). We have expressed the AQP1 protein in yeast secretory vesicles and have also reconstituted yeast-expressed AQP1 into lipid vesicles. We have shown that the protein acts as a water channel in both environments. We are in the process of expressing AQP2 and AQP3 in the same yeast system. We have expressed two closely related proteins (FPS and GlpF) from the MIP family which, although homologous in sequence, do not act as water channels. We would like to find specific amino acids or protein domains which differ among the water channels and non-water channels and which may be important for determining water permeability. To do this, we plan to analyze the sequences and secondary structures of all of the MIP and AQP proteins and look for regions which are conserved or variable. Sites which potentially determine water permeability can then be evaluated experimentally using our yeast expression system. Sequence alignments and secondary structure predictions are possible with the CGC software package which is available from the Pittsburgh Supercomputing Center.