SaccuFlow: A High-Throughput Analysis Platform to Investigate Bacterial Cell Wall Interactions,ACS Infectious Diseases

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SaccuFlow: A High-Throughput Analysis Platform to Investigate Bacterial Cell Wall Interactions
ACS Infectious Diseases ( IF 4.0 ) Pub Date : 2021-07-22 , DOI: 10.1021/acsinfecdis.1c00255
Alexis J Apostolos ₁ , Noel J Ferraro ₁ , Brianna E Dalesandro ₁ , Marcos M Pires ₁

Affiliation

Bacterial cell walls are formidable barriers that protect bacterial cells against external insults and oppose internal turgor pressure. While cell wall composition is variable across species, peptidoglycan is the principal component of all cell walls. Peptidoglycan is a mesh-like scaffold composed of cross-linked strands that can be heavily decorated with anchored proteins. The biosynthesis and remodeling of peptidoglycan must be tightly regulated by cells because disruption to this biomacromolecule is lethal. This essentiality is exploited by the human innate immune system in resisting colonization and by a number of clinically relevant antibiotics that target peptidoglycan biosynthesis. Evaluation of molecules or proteins that interact with peptidoglycan can be a complicated and, typically, qualitative effort. We have developed a novel assay platform (SaccuFlow) that preserves the native structure of bacterial peptidoglycan and is compatible with high-throughput flow cytometry analysis. We show that the assay is facile and versatile as demonstrated by its compatibility with sacculi from Gram-positive bacteria, Gram-negative bacteria, and mycobacteria. Finally, we highlight the utility of this assay to assess the activity of sortase A from Staphylococcus aureus against potential antivirulence agents.

中文翻译：

SaccuFlow：研究细菌细胞壁相互作用的高通量分析平台

细菌细胞壁是强大的屏障，可保护细菌细胞免受外部侵害并抵抗内部膨胀压力。虽然细胞壁组成因物种而异，但肽聚糖是所有细胞壁的主要成分。肽聚糖是一种网状支架，由交联链组成，可以用锚定蛋白进行大量装饰。肽聚糖的生物合成和重塑必须受到细胞的严格调控，因为对这种生物大分子的破坏是致命的。人类先天免疫系统利用这种必要性来抵抗定植，以及许多针对肽聚糖生物合成的临床相关抗生素。评估与肽聚糖相互作用的分子或蛋白质可能是一项复杂的工作，通常是定性的工作。我们开发了一种新型检测平台 (SaccuFlow)，可保留细菌肽聚糖的天然结构，并与高通量流式细胞术分析兼容。我们表明该检测方法简便且用途广泛，正如其与革兰氏阳性菌、革兰氏阴性菌和分枝杆菌的球囊的兼容性所证明的那样。最后，我们强调了该测定在评估分选酶 A 活性方面的效用金黄色葡萄球菌对抗潜在的抗毒剂。

更新日期：2021-08-13

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