套索肽microcinJ25(MccJ25)具有很强的抗菌性能,被认为是细菌性疾病治疗药物和安全食品防腐剂的潜在有效成分。尽管MccJ25可以在枯草芽孢杆菌中异源表达,正如我们以前报道的那样,它的调节和积累还有待理解。这里,我们研究了MccJ25在肽酶基因pepA中断的枯草芽孢杆菌菌株中的表达水平和稳定性,pepF,还有PepT.发现寡内肽酶F(PepF)通过降解其前体肽而降低了MccJ25的产量。在pepF突变体中,培养时间超过60小时后,MccJ25达到1.68µM的浓度,而野生型菌株的浓度仅为0.14µM。此外,枯草芽孢杆菌中MccJ25的产生下调了与孢子形成相关的基因,这可能有助于它的积累。最后,本研究为提高枯草芽孢杆菌中MccJ25的稳定性和产量提供了策略。
目的:MccJ25具有显著的抗菌活性,明确定义的行动模式,特殊的安全性,和显著的稳定性。因此,它是最佳抗菌或抗内毒素药物的令人信服的候选药物。枯草芽孢杆菌外源生产MccJ25的成功建立为降低其生产成本和多样化利用提供了策略。在这项研究中,我们提供的证据表明,肽酶PepF和孢子形成是限制MccJ25在枯草芽孢杆菌中表达的重要因素。枯草芽孢杆菌的ΔpepF和ΔsigF突变体以更高的产量和增强的稳定性表达MccJ25。总而言之,这项研究开发了几个更好的工程菌株枯草芽孢杆菌,这大大减少了在宿主菌株的营养耗尽阶段MccJ25的消耗,提高了产量,并阐明了可能参与减少枯草芽孢杆菌中MccJ25积累的因素。
The lasso peptide microcin J25 (MccJ25) possesses strong antibacterial properties and is considered a potential effective component of bacterial disease treatment drugs and safe food preservatives. Although MccJ25 can be heterologously expressed in Bacillus subtilis as we have previously reported, its regulation and accumulation are yet to be understood. Here, we investigated the expression level and stability of MccJ25 in B. subtilis strains with disruption in peptidase genes pepA, pepF, and pepT. Oligoendopeptidase F (PepF) was found to be involved in reduction of the production of MccJ25 by degradation of its precursor peptide. In the pepF mutant, the MccJ25 reached a concentration of 1.68 µM after a cultivation time exceeding 60 hours, while the wild-type strain exhibited a concentration of only 0.14 µM. Moreover, the production of MccJ25 in B. subtilis downregulated the genes associated with sporulation, and this may contribute to its accumulation. Finally, this study provides a strategy to improve the stability and production of MccJ25 in B. subtilis.
OBJECTIVE: MccJ25 displays significant antibacterial activity, a well-defined mode of action, exceptional safety, and remarkable stability. Hence, it presents itself as a compelling candidate for an optimal antibacterial or anti-endotoxin medication. The successful establishment of exogenous production of MccJ25 in Bacillus subtilis provides a strategy for reducing its production cost and diversifying its utilization. In this study, we have provided evidence indicating that both peptidase PepF and sporulation are significant factors that limit the expression of MccJ25 in B. subtilis. The ΔpepF and ΔsigF mutants of B. subtilis express MccJ25 with higher production yield and enhanced stability. To sum up, this study developed several better engineered strains of B. subtilis, which greatly reduced the consumption of MccJ25 during the nutrient depletion stage of the host strain, improved its production, and elucidated factors that may be involved in reducing MccJ25 accumulation in B. subtilis.