RUI: Enzyme Characterization and Gene Cloning of a Putative Drosophila Homolog for Mammalian Dipeptidyl Peptidase IV

脯氨酸特异性的N-末端加工对于许多生物学上重要的蛋白质和肽的活化和/或降解都是至关重要的。N-末端脯氨酰二肽基肽酶（DPP IV）已在许多生物体中被鉴定.这些包括几种哺乳动物，从小鼠到人，青蛙，真菌，线虫，苍蝇，海葵和几种细菌。 DPP IV在多肽的肠消化和肾转运期间参与肽的代谢。它在哺乳动物免疫系统中活化T淋巴细胞，以及调节造血系统中的DNA合成、细胞增殖和细胞因子产生中发挥作用。 在一些哺乳动物中，它在信号转导和细胞粘附现象中起作用。 在人类神经系统中，它调节神经肽Y的活性，并且在海葵中可能在至少一种神经肽（变态蛋白A）的加工中发挥作用。在昆虫中，DPP IV被认为在几种肽的加工中起作用，包括几种物种的免疫肽和蜜蜂毒液中的肽蜂毒肽。然而，只有少数天然底物的酶已被证实。 该实验室最近报道了果蝇中的突变体（omega），该突变体已经失去了DPP IV酶，因此显示出几种亚致死缺陷：幼虫发育延迟，雄性生育力下降40%，雄性成功使雌性受精的能力下降，以及幼虫角质层蛋白翻译后加工的损失。 这些缺陷表明果蝇中需要这种酶的多种过程，这意味着除了角质层蛋白外，还有几种未知的肽被DPP IV加工。 该研究包括计划利用伯克利果蝇基因组计划的现有基因组克隆从果蝇中克隆DPP IV基因，其中一个基因组克隆最近被鉴定为具有嵌入其中的同源序列，这将允许将其结构与已知的来自其他生物的DPP IV基因进行比较。酶也将被纯化。这将允许表征其生化特性。 DPP IV在果蝇肽加工中的作用，特别是在角质层蛋白、免疫肽和睾丸肽方面，将使用可用的注射和PAGE方法进行澄清。 这些数据将导致进一步了解DPP IV在先天免疫系统和果蝇的其他肽加工系统中的功能，从而进一步证明这种重要酶在真核生物中的普遍性。

Proline specific N-terminal processing is vital to both the activation and/or degradation of many biologically important proteins and peptides. The N- terminal prolyl dipeptidyl peptidase (DPP IV ) has been identified in an impressive array of organisms. These include several species of mammal, from mouse to man, frog, fungi, nematode, flies, sea anemone, and several species of bacteria. DPP IV is involved in the metabolism of peptides during the intestinal digestion and renal transport of polypeptides. It functions in the activation of T-lymphocytes in the immune system of mammals, and the regulation of DNA synthesis, cell proliferation and cytokine production in the hematopoietic system. In some mammals, it functions in signal transduction and cell adherence phenomena. In the human nervous system, it regulates the activity of neuropeptide Y and in the sea anemone is likely to play a role in the processing of at least one neuropeptide (metamorphosin A). In insects, DPP IV is presumed to function in the processing of several peptides, including the immune peptides of several species and the peptide melittin in honeybee venom. However, only a few natural substrates for the enzyme have been confirmed. This laboratory recently reported on a mutant stock (omega) in the fruit fly Drosophila melanogaster that has lost the enzyme DPP IV and as a consequence displays several sublethal defects: A larval developmental delay, a 40% decrease in male fertility and in the ability of males to successfully fertilize females, and a loss of post translational processing of a larval cuticle protein. These defects point to a multiplicity of processes for which the enzyme is required in Drosophila, implying that there are several, as yet unknown, peptides that are processed by DPP IV in addition to the cuticle protein. This research includes plans to clone the DPP IV gene from Drosophila using the available genomic clones of the Berkeley Drosophila Genome Project, one of which has recently been identified as having an homologous sequence imbedded in it. This will allow comparison of its structure with the known DPP IV genes from other organisms. The enzyme will also be purefied. This will allow characterization of its biochemical properties. The role of DPP IV in the processing of Drosophila peptides, in particular with respect to the cuticle proteins, the immune peptides and testicular peptides, will be clarified using available injection and PAGE procedures. These data will lead to a further understanding of the DPP IV function in the innate immune system and in other peptide processing systems of Drosophila, thus further demonstrating the ubiquity of this important enzyme in eukaryotic organisms.