Effect of PSMα knockout on autolysin AtlA expression in <italic style="font-style: italic">Staphylococcus aureus</italic>

Li TAN; Sirui LI; Bei JIANG; Xiaopeng ZHANG; Jiang CHEN; Xiaomei HU; Shu LI

doi:10.16016/j.1000-5404.201706121

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Effect of PSMα knockout on autolysin AtlA expression in Staphylococcus aureus

Basic Medicine | 更新时间：2026-01-09

- Effect of PSMα knockout on autolysin AtlA expression in Staphylococcus aureus
- Journal of Third Military Medical University Vol. 39, Issue 21, Pages: 2066-2071(2017)
- 作者机构：
  
  400038 重庆，第三军医大学基础医学部微生物学教研室
- 作者简介：
- 基金信息：
  
  the National Natural Science Foundation of China(31470241);the National Natural Science Foundation of China(31570127);the Innovation and Entrepreneurship Training Program for Undergraduates in China(201690031002)
- DOI：10.16016/j.1000-5404.201706121
  CLC： R378.11;R394-33;R394.3
- Received：21 June 2017，
  
  Revised：2017-07-21，
  
  Published：15 November 2017
- 稿件说明：
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Li TAN, Sirui LI, Bei JIANG, et al. Effect of PSMα knockout on autolysin AtlA expression in Staphylococcus aureus[J]. Journal of Third Military Medical University, 2017, 39(21): 2066-2071.
DOI：

Li TAN, Sirui LI, Bei JIANG, et al. Effect of PSMα knockout on autolysin AtlA expression in Staphylococcus aureus[J]. Journal of Third Military Medical University, 2017, 39(21): 2066-2071. DOI： 10.16016/j.1000-5404.201706121.

摘要

目的

利用同源重组技术构建金黄色葡萄球菌Newman psmα基因缺失突变株，探究psmα缺失对金黄色葡萄球菌蛋白表达的影响。

方法

分别扩增psmα基因的上下游同源臂，根据同源重组原理，构建pBT2基因打靶载体，利用同源重组技术，构建psmα缺失的Newman突变株。并对野生株和突变株培养上清蛋白进行研究。

结果

突变株培养上清中一个相对分子质量约为135×10

的蛋白表达水平较野生株明显减少甚至缺失；质谱分析提示该蛋白为金黄色葡萄球菌atlA基因编码的双功能自溶素前体蛋白(AtlA蛋白)；实时荧光定量PCR结果显示psmα基因敲除株atlA基因的转录水平较野生株显著降低。

结论

psmα缺失后能够降低atlA基因的转录表达水平。

Abstract

Objective

To construct the phenol-soluble modulin (psmα) locus gene knockout mutant of

Staphylococcus aureus

(

S. aureus

) and evaluate the effect of PSMα peptides on the expression of secretory proteins.

Methods

The temperature-sensitive

S. aureus

knock-out vectors were constructed after 2 homologous sequences around the psmα locus were cloned and inserted into pBT2 vector to construct a knockout plasmid pBT2∷psmα. Then the obtained plasmids pBT2∷psmα were transferred into

S. aureus

strain Newman

and the Newman∷Δpsmα mutant were prepared with homologous recombination. The proteins in the culture supernatants of Newman wild type strain and Newman∷Δpsmα mutant were analyzed with SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry. The transcriptional level of altA gene was detected using quantitative real-time PCR.

Results

SDS-PAGE confirmed that there was a band missing at the relative mass weight of 135×10

when compared with the results of the wildtype strain. Mass spectrometry indicated that the protein was dual functionality autolyzed precursor protein (AtlA) in Newman∷Δpsmα mutant. RT-PCR also showed the atlA gene transcription was significantly reduced in the psmα mutant strain.

Conclusion

The PSMα knockout in S. aureus reduces the expression of atlA at transcriptional level.

关键词

Keywords

references

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Effect of PSMα knockout on autolysin AtlA expression in Staphylococcus aureus

DOI：10.16016/j.1000-5404.201706121

摘要

Abstract

关键词

Keywords

references