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首页> 外文期刊>Virulence >Determination of essentiality and regulatory function of staphylococcal YeaZ in branched-chain amino acid biosynthesis
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Determination of essentiality and regulatory function of staphylococcal YeaZ in branched-chain amino acid biosynthesis

机译：确定葡萄球菌YeaZ在支链氨基酸生物合成中的必要性和调节功能

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The staphylococcal YeaZ is highly conserved in prokaryotic cells and critical for growth of many bacterial pathogens. However, the essentiality for Staphylococcus aureus growth and the biological function of YeaZ behind its essentiality remain undefined. In this study, we created and characterized a defined Pspac-regulated yeaZ expression mutant in S. aureus and demonstrated the indispensability of YeaZ for S. aureus growth. Moreover, we conducted complementation studies, not only confirmed the requirement of YeaZ for S. aureus growth, but also revealed a similarity of essential function between staphylococcal YeaZ and its E. coli homolog. On the other hand, we explored the biological functions of YeaZ and found that YeaZ is involved in the regulation of the transcription of ilv-leu operon that encodes key enzymes responsible for the biosynthesis of the branched-chain amino acids, including isoleucine, leucine, and valine (ILV). qPCR analysis showed that the 6-fold downregulation of YeaZ dramatically elevated approximately 17- to 289-fold RNA levels of ilvD, leuA and ilvA. We further confirmed the transcriptional regulation of the ilv-leu operon by YeaZ using an ilv-promoter-lux reporter system and gel-shift assays and revealed that YeaZ is able to bind the promoter region of ilv. Furthermore, we established that the regulation of ILV biosynthesis isn't associated with YeaZ's essentiality, as the deletion of the ilv-leu operon did not affect the requirement of YeaZ for growth in culture. Our results demonstrate the essentiality of YeaZ for S. aureus growth and suggest that the staphylococcal YeaZ possesses regulatory function.
机译：葡萄球菌YeaZ在原核细胞中高度保守，对许多细菌性病原体的生长至关重要。然而，金黄色葡萄球菌生长的必要性以及其必需性背后的YeaZ生物学功能仍然不确定。在这项研究中，我们创建并表征了金黄色葡萄球菌中Pspac调控的yeaZ表达突变体，并证明了YeaZ对于金黄色葡萄球菌生长不可或缺。此外，我们进行了互补研究，不仅证实了YeaZ对金黄色葡萄球菌生长的需求，而且还揭示了葡萄球菌YeaZ和其大肠杆菌同源物之间基本功能的相似性。另一方面，我们探索了YeaZ的生物学功能，并发现YeaZ参与了ilv-leu操纵子转录的调控，该操纵子编码负责支链氨基酸生物合成的关键酶，包括异亮氨酸，亮氨酸，和缬氨酸（ILV）。 qPCR分析表明，YeaZ的6倍下调显着提高了ilvD，leuA和ilvA的17到289倍RNA水平。我们进一步证实了使用ilv-promoter-lux报告基因系统和凝胶位移测定法通过YeaZ对ilv-leu操纵子的转录调控，并揭示了YeaZ能够结合ilv的启动子区域。此外，我们确定ILV生物合成的调控与YeaZ的必要性无关，因为ilv-leu操纵子的缺失并不影响YeaZ对培养物生长的需求。我们的结果证明了YeaZ对于金黄色葡萄球菌生长的必要性，并表明葡萄球菌YeaZ具有调节功能。

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