转录调控因子在pqqA转录激活和PQQ快速合成过程中的应答机制

项目名称： 转录调控因子在pqqA转录激活和PQQ快速合成过程中的应答机制

项目编号： No.31300083

项目类型： 青年科学基金项目

立项/批准年度： 2014

项目学科： 生物科学

项目作者： 葛欣

作者单位： 中国人民解放军军事医学科学院

项目金额： 23万元

中文摘要： 吡咯喹啉醌（Pyrroloquinoline quinone,PQQ）是细菌的第三类氧化还原辅酶，具有重要的生理功能和药用开发价值。Methylovorus sp.MP688是极具产业化前景的PQQ生产菌，基因簇pqqABCDE是PQQ合成的遗传基础，其中pqqA编码的小蛋白是PQQ合成的直接底物。实验发现PQQ高效合成的前提是pqqA的高效表达；而细菌生长时pqqA只维持着基础转录，在复杂的胞外信号如低pH、低溶氧、低钙等条件下才能被高效诱导；启动子截短发现了一段抑制pqqA转录的DNA区域，进而通过生化分离和遗传学手段鉴定了转录调控蛋白。本研究通过比较蛋白组学、蛋白质相互作用、分子遗传学等方法考察转录调控因子特征及在胞外信号作用下调节基因转录的方式和差别，阐明其在pqqA转录激活和PQQ快速合成过程中的作用机制，为构建pqqA基因持续高效转录的菌株和最终提高PQQ水平提供理论依据。

中文关键词： 吡咯喹啉醌；甲基营养菌；转录因子；转录组；

英文摘要： Pyrroloquinoline quinone is the third class of coenzyme and a newly discovered vitamin, which exerts physiological functions in prevention and cure hepatic injury, promoting growth of NGF, balancing the free radical, protecting cells from radiation injuries. As a result, PQQ is a promising candidate drug in therapy. Methylovorus sp. MP688 can syntheisi a relative high leve of PQQ. A small peptide PqqA encoded by gene pqqA is the precursor and substrate of PQQ. Experiment results indicate that large-scale production of PQQ requires a high lever transcription of pqqA gene. And pqqA was effectively induced in lower pH, lower dissolval oxygen and calcium concentration. Deletion analysis revealed a DNA region inhibiting pqqA transcription. After that, putative transcriptional regulators were isolated by biochemical and genetic method. This research focus on the transcription regulator, either positive or negative, of pqqA gene by using comparative genomics, protein-protein interaction and genetic methods. Furthermore, we study how the regulator response to the signals, causing the signal transduction and effecting at the target DNA sequences. On the basis of elucidating the transcriptional regulation mechanism of pqqA gene, we will make some improvement of the PQQ-producing strain in order to enhance the production.

英文关键词： Pyrroloquinoline quinone；Methylotrophic bacteria；Transcription factor；Transcriptome；