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

研究了影响枯草芽孢杆菌NAFM5胞外谷氨酸多聚体--聚γ-谷氨酸(PGA)合成和降解的营养因素。我们还分析了参与PGA生物合成和降解酶基因调控的调控系统。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节：L-谷氨酸和葡萄糖分别促进PGA的合成2.5倍和3倍，而CAPB：：LacZ融合的表达分别被抑制到1/2和1/6。结果表明：1.含有大豆蛋白水解物的营养丰富的GSP培养基能显著提高(是微量培养基中的11倍)，而LB培养基对CAPB：：LacZ的表达和PGA的产生几乎没有抑制作用。通过群体感应系统对capBCAE PGA生物合成操纵子的调节，我们发现des Q和des SU基因是PGA合成所必需的，并通过凝胶位移分析确定deGU与CAPB启动子的-720 ro-660区域结合。YwtD和GGT的调节。]我们发现GGT受ComQXPA群体感应系统控制。遗传学研究表明，ywrD编码一种终止型PGA水解酶。该基因似乎不受群体感应系统的调控。VvzD的功能分析利用yvzD的敲除突变体，我们证明了该基因编码一种与细胞壁分离相关的脱锚酶PGA。因此，至少有三个基因，yvzD，ywtD和ggt，参与了PGA的降解。YvzD也不受法定人数感应系统的控制。

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.