Chaussee Michelle A, McDowell Emily J, Rieck Lindsey D, Callegari Eduardo A, Chaussee Michael S
Division of Basic Biomedical Sciences, The Stanford School of Medicine of the University of South Dakota 414 East Clark Street, Vermillion, SD 57069, USA.
J Antimicrob Chemother. 2006 Oct;58(4):752-9. doi: 10.1093/jac/dkl319. Epub 2006 Aug 5.
To determine whether the transcriptional regulator Rgg contributes to penicillin-induced killing of Streptococcus pyogenes by altering a regulatory response to penicillin.
Penicillin-induced killing of a wild-type and isogenic rgg mutant strain was assessed in broth and solid media and in the presence of cerulenin, which inhibits fatty acid biosynthesis (FAB). Proteins from wild-type and rgg mutant cultures, either exposed to penicillin or not, were characterized by two-dimensional gel electrophoresis. Proteins of interest were identified with tandem mass spectrometry.
The MIC of penicillin was 0.012 mg/L for both the wild-type strain NZ131 and an isogenic rgg mutant strain. The wild-type strain lost 1.9 log(10) cfu/mL ( approximately 80-fold) after 24 h of exposure to 0.024 mg/L penicillin compared with controls; however, the mutant strain lost 0.3 log(10) cfu/mL ( approximately 2-fold) compared with controls. Changes in the proteome of wild-type and mutant cultures were assessed 1 and 4 h after exposure to penicillin. One hour exposure was associated with increased abundance (P < 0.05) of 12 proteins associated with FAB, the pentose phosphate pathway, glycolysis and stress responses in the wild-type strain. The abundance of 8 of 12 of these proteins was greater in samples obtained from the mutant strain, even prior to penicillin exposure. After 4 h of exposure, the abundance of 16 proteins was altered in one or both strains; however, a clear functional relationship was not evident. The addition of cerulenin slightly enhanced penicillin-induced killing of wild-type strain, which supported the proteomic results.
The results suggest that penicillin-independent changes in the cytoplasmic proteome of an rgg mutant strain of NZ131 confer tolerance to penicillin-mediated killing.
通过改变对青霉素的调节反应，确定转录调节因子Rgg是否有助于青霉素诱导的化脓性链球菌杀伤作用。
在肉汤和固体培养基中以及在存在抑制脂肪酸生物合成（FAB）的铜绿霉素的情况下，评估青霉素对野生型和同基因rgg突变株的杀伤作用。通过二维凝胶电泳对野生型和rgg突变株培养物中未接触或接触青霉素的蛋白质进行表征。感兴趣的蛋白质通过串联质谱法进行鉴定。
野生型菌株NZ131和同基因rgg突变株的青霉素最低抑菌浓度（MIC）均为0.012 mg/L。与对照相比，野生型菌株在接触0.024 mg/L青霉素24小时后，cfu/mL损失了1.9 log(10)（约80倍）；然而，突变株与对照相比，cfu/mL损失了0.3 log(10)（约2倍）。在接触青霉素1小时和4小时后，评估野生型和突变株培养物蛋白质组的变化。接触1小时与野生型菌株中12种与FAB、磷酸戊糖途径、糖酵解和应激反应相关的蛋白质丰度增加（P < 0.05）有关。在从突变株获得的样品中，甚至在接触青霉素之前，这12种蛋白质中的8种丰度更高。接触4小时后，16种蛋白质的丰度在一个或两个菌株中发生了变化；然而，明显的功能关系并不明显。添加铜绿霉素略微增强了青霉素对野生型菌株的杀伤作用，这支持了蛋白质组学结果。
结果表明，NZ131的rgg突变株细胞质蛋白质组中与青霉素无关的变化赋予了对青霉素介导杀伤的耐受性。
