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The mechanism of the beta-lactam resistance in Staphylococcus aureus

金黄色葡萄球菌β-内酰胺耐药机制

基本信息

批准号：
10674849
负责人：
Taeok Bae
金额：
$ 39.63万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-01 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10674849
关键词：
ATP-Dependent Proteases Affect Anti-Bacterial Agents Antibiotic Resistance Antibiotics Bacteria Bacterial Infections Binding Binding Proteins Biological Assay Cell Wall Cell membrane Chromosomes Development Drug resistance Enzymes Genes Genome Goals Infection Lactams Mediating Membrane Methicillin Resistance Molecular Mutagenesis Mutation Pathway interactions Penicillin-Binding Proteins Peptide Hydrolases Peptidoglycan Pharmaceutical Preparations Pharmacotherapy Phenotype Pneumonia Production Resistance Resistance development Role Sepsis Skin Tissue Soft Tissue Infections Staphylococcal Infections Staphylococcus aureus Staphylococcus aureus infection Suppressor Genes Suppressor Mutations Testing Therapeutic Agents Toxic Shock Syndrome beta-Lactam Resistance beta-Lactams coping design enzyme activity genome sequencing infectious disease treatment lipoteichoic acid methicillin resistant Staphylococcus aureus minimal risk mutant novel novel therapeutics overexpression pathogenic bacteria protein function resistance mechanism resistance mutation resistant strain skin disorder transposon sequencing yeast two hybrid system

项目摘要

Project Summary Cell-wall targeting beta-lactam antibiotics are the largest group of antibacterial agents and have been the mainstay for treatment of bacterial infections. However, most beta-lactam antibiotics cannot be used for the treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections because MRSA produces a cell- wall synthesis enzyme PBP2A, which most beta-lactams cannot inactivate. Although PBP2A is the primary determinant of MRSA beta-lactam resistance, the overall beta-lactam resistance of MRSA is affected by additional bacterial factors such as FtsH, a protease in the cell membrane. Overexpression of FtsH makes MRSA sensitive specifically to beta-lactams. The beta-lactam-sensitizing effect of FtsH is due to its degradation of YpfP, an enzyme synthesizing the anchor molecule for lipoteichoic acid (LTA). The YpfP- degradation by FtsH causes the production of abnormally large LTA. On the other hand, increased production of normal LTA by deletion of the ftsH gene makes MRSA more resistant to beta-lactams. When the production of normal LTA is suppressed by inhibiting the production of the LTA synthesis enzyme LtaS, MRSA becomes hypersensitive to beta-lactams. Based on these results, this study hypothesizes that the large LTA lowers MRSA beta-lactam resistance by suppressing the enzyme activities of the cell-wall synthesis enzymes whereas the normal LTA is required for them. Even under the beta-lactam sensitizing conditions, MRSA can regain resistance to beta-lactams. Genome-sequencing of 26 such mutants showed that 16 genes are critical for MRSA beta-lactam resistance. The long-term goal of this project is to develop novel β-lactam potentiators against MRSA. The objective of this study is to understand how FtsH and the disturbance in LTA synthesis sensitize MRSA to β-lactams, and how MRSA regains β-lactam resistance. In aim 1, with the MRSA strains producing either the large LTA or reduced amount of the normal LTA, the cell wall synthesis steps affected by the LTA molecules will be determined. The effect of LTA on cell wall synthesis will be directly tested by a peptidoglycan-synthesis assay. Also, the bacterial two-hybrid analysis will determine whether the degradation of YpfP is modulated by FtsH-binding proteins. In aim 2, by generating each of the resistance mutations in the chromosome of MRSA, the mutations critical for MRSA β-lactam resistance will be determined. Also, transposon-mediated mutagenesis will identify the genes essential for MRSA to regain the β-lactam resistance. Finally, to test whether the roles of the 16 genes are universally conserved among S. aureus strains, resistant mutants will be identified from two more MRSA strains, and their genomes will be sequenced. Completion of this study will reveal the intricate network of the beta-lactam resistance regulators in MRSA and open the door to developing novel beta-lactam potentiators with minimal risk of resistance development in the treatment of MRSA infections. Such drugs are expected to reduce the burdens of antibiotic resistance.

项目摘要细胞壁靶向β-内酰胺抗生素是最大的一类抗菌剂，是治疗细菌感染的主要药物。然而，大多数β-内酰胺抗生素不能用于治疗耐甲氧西林金黄色葡萄球菌（MRSA）感染，因为MRSA产生一种细胞- 壁合成酶PBP 2A，大多数β-内酰胺不能降解。虽然PBP 2A是主要的作为MRSA β-内酰胺耐药性的决定因素，MRSA的总体β-内酰胺耐药性受以下因素的影响：另外的细菌因子如FtsH，细胞膜中的蛋白酶。FtsH的过表达使得对β-内酰胺类药物敏感的MRSA。FtsH的β-内酰胺致敏作用是由于其 YpfP的降解，YpfP是合成脂磷壁酸（LTA）的锚分子的酶。YpfP- FtsH的降解导致产生异常大的LTA。另一方面，产量增加通过缺失ftsH基因使正常的LTA减少，使MRSA对β-内酰胺类抗生素更具耐药性。当生产通过抑制LTA合成酶LtaS的产生来抑制正常LTA的合成，MRSA成为对β-内酰胺过敏基于这些结果，本研究假设，大的LTA降低了通过抑制细胞壁合成酶的酶活性来产生MRSA β-内酰胺耐药性而正常的LTA是他们所需要的。即使在β-内酰胺致敏条件下，MRSA也可以恢复对β-内酰胺类药物的耐药性。对26个突变体的基因组测序表明，16个基因是关键的耐甲氧西林金黄色葡萄球菌β-内酰胺耐药性本项目的长期目标是开发新型β-内酰胺增效剂抗MRSA本研究的目的是了解FtsH和干扰在LTA合成中的作用使MRSA对β-内酰胺类药物敏感，以及MRSA如何恢复β-内酰胺类药物耐药性。在目标1中，使用MRSA菌株产生大的LTA或减少量的正常LTA，细胞壁合成步骤受将确定LTA分子。LTA对细胞壁合成的影响将通过一个直接测试。肽聚糖合成测定。此外，细菌双杂交分析将确定降解是否 YpfP的表达受FtsH结合蛋白的调节。在目标2中，通过在细胞中产生每种抗性突变，在MRSA的染色体上，将确定对MRSA β-内酰胺耐药至关重要的突变。还有，转座子介导的突变将鉴定MRSA恢复β-内酰胺抗性所必需的基因。最后，为了验证这16个基因的功能是否在S.金黄色葡萄球菌菌株，耐药将从另外两种MRSA菌株中鉴定出突变株，并对它们的基因组进行测序。完成这项研究将揭示MRSA中β-内酰胺耐药调节因子的复杂网络，本发明涉及开发新的β-内酰胺增效剂， MRSA感染。这些药物有望减少抗生素耐药性的负担。

项目成果

期刊论文数量（4）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Ftsh Sensitizes Methicillin-Resistant Staphylococcus aureus to β-Lactam Antibiotics by Degrading YpfP, a Lipoteichoic Acid Synthesis Enzyme.

DOI：
10.3390/antibiotics10101198
发表时间：
2021-10-01
期刊：
Antibiotics (Basel, Switzerland)
影响因子：
0
作者：
Yeo WS;Jeong B;Ullah N;Shah MA;Ali A;Kim KK;Bae T
通讯作者：
Bae T

Molybdopterin biosynthesis pathway contributes to the regulation of SaeRS two-component system by ClpP in Staphylococcus aureus.

DOI：
10.1080/21505594.2022.2065961
发表时间：
2022-12
期刊：
VIRULENCE
影响因子：
5.2
作者：
Zhao, Na;Wang, Yanan;Liu, Junlan;Yang, Ziyu;Jian, Ying;Wang, Hua;Ahmed, Mahmoud;Li, Min;Bae, Taeok;Liu, Qian
通讯作者：
Liu, Qian

Unravelling the physiological roles of mazEF toxin-antitoxin system on clinical MRSA strain by CRISPR RNA-guided cytidine deaminase.