A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky pr. 33, Moscow, 119071, Russia.
Antonie Van Leeuwenhoek. 2013 Jan;103(1):37-46. doi: 10.1007/s10482-012-9784-1. Epub 2012 Aug 4.
Resuscitation promoting factors (Rpfs), belonging to a family of secreted actinobacterial proteins with predicted peptidoglycan (PG) hydrolytic activities, participate in the reactivation of dormant cells. In the present study we demonstrate that a recombinant truncated form of Micrococcus luteus Rpf hydrolyzes isolated PG of Mycobacterium smegmatis and Mycobacterium tuberculosis liberating PG fragments of different size. These fragments possess stimulatory activity toward "non-culturable" dormant M. smegmatis and M. tuberculosis cells, similar to the activity of recombinant Rpf. Relatively large PG fragments (0.1-0.5 μm) obtained either by Rpf digestion or by PG ultrasonication revealed resuscitation activities when added in concentrations 0.1-0.2 μg/ml to the resuscitation medium. It is suggested that PG fragments could either directly activate the resuscitation pathway of dormant mycobacteria or serve as a substrate for endogenous Rpf, resulting in low molecular weight products with resuscitation activity. Whilst both suggestions are plausible, it was observed that PG-dependent resuscitation activity was suppressed by means of a specific Rpf inhibitor (4-benzoyl-2-nitrophenylthiocyanate), which provides additional support for the second of these possibilities.
复苏促进因子（Rpfs）属于一类具有预测肽聚糖（PG）水解活性的分泌放线菌蛋白家族，参与休眠细胞的再激活。在本研究中，我们证明了一种来自微球菌的重组截断形式的 Rpf 可以水解分枝杆菌属的分离 PG，释放出不同大小的 PG 片段。这些片段对“非可培养”的休眠分枝杆菌和结核分枝杆菌细胞具有刺激活性，类似于重组 Rpf 的活性。通过 Rpf 消化或 PG 超声获得的相对较大的 PG 片段（0.1-0.5μm），当添加到再培养液中的浓度为 0.1-0.2μg/ml 时，具有再激活活性。这表明 PG 片段可以直接激活休眠分枝杆菌的再激活途径，或者作为内源性 Rpf 的底物，产生具有再激活活性的低分子量产物。虽然这两种假设都有一定的合理性，但我们观察到 PG 依赖性再激活活性被一种特定的 Rpf 抑制剂（4-苯甲酰基-2-硝基苯硫氰酸酯）抑制，这为第二种可能性提供了额外的支持。
