Studies on the enzyme that catalyzes D/L-isomerization of an amino acid during the post-translational modification of neuropeptide precursor.

NdWFamide （NdWFa）是一种含有d氨基酸的海洋腹足类动物黑黑的神经肽。在这项研究中，我们试图鉴定在美国催化NdWFa的D-/ l转化的肽基D/ l异构酶（PI）。为此，我们采用减法制备了表达ndwfa的神经元特异性cdna。在对这些dna进行克隆和测序时，我们发现一些带有n端信号肽的预测蛋白无法通过对数据库的同源性搜索确定其功能。目前，我们认为这些蛋白是蓝叶菊PI的候选蛋白。然后，我们尝试建立表达ndwfa的细胞系，因为这样的细胞可以用于PI候选基因的生物测定系统。ndwfa前体基因向CHO-K1细胞的转化似乎是成功的。然而，到目前为止，该肽的生物合成尚未得到证实。我们可能需要微调实验条件，从而更有效地表达外源基因。

NdWFamide (NdWFa) is a D-amino acid containing neuropeptide of the marine gastropod, Aplysia kurodai. In this study, we attempt to identify the peptidyl D/L-isomerase (PI) that catalyzes the D-/L-conversion of NdWFa in Aplysia. To this end, we prepared NdWFa-expressing neuron specific cDNAs by the subtraction method. When the cDNAs were cloned and sequenced, we found that several predicted proteins with N-terminal signal peptide could not be specified their functions by homology search on database. We currently assume that those proteins are candidates for the PI of Aplysia.Then, we tried to establish NdWFa-expressing cell-line, because such cells could be used for the bioassay system for the candidate genes of the PI. Transformation of NdWFa-precursor gene to CHO-K1 cell seems to be successful. However, biosynthesis of the peptide could not be confirmed, so far. We may need fine-tuning of experimental conditions that leads to more effective expression of foreign gene.