Functional analyses of bacterial nucleotide transport proteins

利用宿主衍生的核苷酸和“能量寄生”是某些专性细胞内（致病）细菌的基本共同特征。尽管迄今为止各种核苷酸转运蛋白（NTT）已被表征，但有关其生化特性和结构/功能关系的基本问题仍未得到解决。我们最近的分析提供了磷酸盐作为ATP/ADP转运体PamNTT1的先前未被识别的底物的证据，这些转运体来自变形虫共栖原衣原体变形虫。有趣的是，NTT驱动的磷酸盐运输将使专门的磷酸盐载体变得不必要，这也解释了为什么在细胞内活细菌中寻找相应蛋白质的长期努力失败了。在这项研究中，我们将对功能不同的核苷酸转运体PamNTT1-5和立克次氏体中具有代表性的ATP/ADP转运体进行复杂的生化分析，目的是澄清细菌磷酸转运是否(i)仅限于PamNTT1或（ii）“能量寄生”的典型特征或（iii）细菌核苷酸转运的一般原则。此外，通过选择突变蛋白的帮助下，我们的目标是鉴定核苷酸和磷酸盐运输所必需的氨基酸。

Exploitation of host derived nucleotides and “energy parasitism” are essential common features of certain obligate intracellular (pathogenic) bacteria. Although various nucleotide transport proteins (NTT) have been characterized so far, fundamental questions concerning biochemical properties and structure/function relationships remained unaddressed. Our very recent analyses provided evidence for phosphate as a previously unidentified substrate of the ATP/ADP transporter PamNTT1 from the amoeba symbiont Protochlamydia amoebophila. Interestingly, an NTT driven phosphate transport would make a specialized phosphate carrier unnecessary and would explain why the long lasting search for the corresponding protein in intracellular living bacteria failed. In this study, we will perform a sophisticated biochemical analysis of the functionally different nucleotide transporters PamNTT1-5 and also of a representative ATP/ADP transporter from Rickettsiales which aims to clarify whether bacterial phosphate transport is (i) restricted to PamNTT1 or (ii) a typical feature of “energy parasitism” or (iii) a general principle in bacterial nucleotide transport. Furthermore, by the help of selected mutant proteins we aim to identify amino acids essential for nucleotide and phosphate transport.