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Regulation of poly-gamma-glutamate synthesis and degradation in Bacillus subtlis

枯草芽孢杆菌中聚γ-谷氨酸合成和降解的调控

基本信息

批准号：
17380057
负责人：
ITOH Yoshifumi
金额：
$ 7.55万
依托单位：
Tohoku University (2006)Akita Prefectural Agriculture, Forestry and Fisheries Research Center (2005)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

We investigated nutrient factors that affect the synthesis and degradation of extracellular polymer of glutamate, poly-γ-glutamate (PGA), in Bacillus subtilis NAFM5, a derivative of a starter strain for Natto (Japanese fermented soybean food). We also analyzed regulatory systems involved in the regulation of the PGA biosynthetic and degradative enzyme genes. Our major results include :1. Regulation of the capBCAEPGA biosynthetic operon by nutrients.L-Glutamate and glucose stimulated PGA production by 2.5-and 3-fold, but repressed expression of a capB::lacZ fusion to 1/2 and 1/6, respectively. Nutrient-rich GSP medium, which contained soybean protein hydroysates, greatly improved (11 fold of that in minimal medium), whereas LB medium almost prevent capB::lacZ expression and PGA production.2. Regulation of the capBCAE PGA biosynthetic operon by the quorum-sensing system.We found that degQ and degSU genes are essential for PGA synthesis and determined by gel-shift assays that DegU binds to-720 ro-660 region of the capB promoter.3. Regulation of ywtD and ggt.]We found that ggt is under the control of the ComQXPA quorum-sensing system. Genetic study showed that ywrD encodes an end-type of PGA hdrolase. This gene appeared not regulated by the quorum-sensing system.4. Functional analysis of vvzD.Using a knockout mutant of yvzD, we demonstrated that this gene encode a deanchoring enzyme detaching cell-wall associated PGA. Thus, at least three genes, yvzD, ywtD and ggt, are involved in PGA degradation. YvzD is also not controlled by the quorum-sensing system.

我们研究了影响胞外谷氨酸聚合物聚γ-谷氨酸（PGA）在枯草芽孢杆菌（Bacillus subtilis NAFM5）中合成和降解的营养因子。NAFM5是纳豆（日本发酵大豆食品）发酵剂的衍生物。我们还分析了参与调控PGA生物合成和降解酶基因的调控系统。我们的主要结果包括：1。营养物质对capBCAEPGA生物合成操纵子的调控。l -谷氨酸和葡萄糖对PGA产生的刺激分别为2.5倍和3倍，但对capB::lacZ融合表达的抑制分别为1/2和1/6。含有大豆蛋白水解物的富营养GSP培养基显著提高了大豆蛋白水解产物的表达（是最低培养基的11倍），而LB培养基几乎阻止了capB::lacZ的表达和PGA的产生。群体感应系统对capBCAE PGA生物合成操纵子的调控。我们发现degQ和degSU基因在PGA合成中是必需的，并且通过凝胶转移实验确定DegU结合到capB启动子的720 o-660区域。ywtD和ggt的调节。我们发现ggt是在ComQXPA群体感应系统的控制下。遗传研究表明，ywrD编码一种末端型PGA水解酶。该基因似乎不受群体感应系统的调节。vvzD的功能分析。使用yvzD的敲除突变体，我们证明该基因编码一种分离细胞壁相关PGA的去锚定酶。因此，至少有三个基因yvzD、ywtD和ggt参与了PGA的降解。YvzD也不受群体感应系统的控制。