Fluorogenic D-amino acids enable real-time monitoring of peptidoglycan biosynthesis and high-throughput transpeptidation assays.,Nature Chemistry

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Fluorogenic D-amino acids enable real-time monitoring of peptidoglycan biosynthesis and high-throughput transpeptidation assays.
Nature Chemistry ( IF 19.2 ) Pub Date : 2019-02-25 , DOI: 10.1038/s41557-019-0217-x
Yen-Pang Hsu ₁ , Edward Hall _{2,

3} , Garrett Booher ₁ , Brennan Murphy ₂ , Atanas D Radkov _{2,

4} , Jacob Yablonowski ₁ , Caitlyn Mulcahey ₁ , Laura Alvarez ₅ , Felipe Cava ₅ , Yves V Brun _{6,

7} , Erkin Kuru _{1,

8} , Michael S VanNieuwenhze _{1,

2}

Affiliation

Peptidoglycan is an essential cell wall component that maintains the morphology and viability of nearly all bacteria. Its biosynthesis requires periplasmic transpeptidation reactions, which construct peptide crosslinkages between polysaccharide chains to endow mechanical strength. However, tracking the transpeptidation reaction in vivo and in vitro is challenging, mainly due to the lack of efficient, biocompatible probes. Here, we report the design, synthesis and application of rotor-fluorogenic D-amino acids (RfDAAs), enabling real-time, continuous tracking of transpeptidation reactions. These probes allow peptidoglycan biosynthesis to be monitored in real time by visualizing transpeptidase reactions in live cells, as well as real-time activity assays of D,D- and L,D-transpeptidases and sortases in vitro. The unique ability of RfDAAs to become fluorescent when incorporated into peptidoglycan provides a powerful new tool to study peptidoglycan biosynthesis with high temporal resolution and prospectively enable high-throughput screening for inhibitors of peptidoglycan biosynthesis.

中文翻译：

荧光 D-氨基酸能够实时监测肽聚糖的生物合成和高通量转肽化分析。

肽聚糖是维持几乎所有细菌的形态和活力的基本细胞壁成分。它的生物合成需要周质转肽反应，在多糖链之间构建肽交联以赋予机械强度。然而，在体内和体外追踪转肽反应具有挑战性，主要是由于缺乏有效的、生物相容的探针。在这里，我们报告了转子荧光 D-氨基酸 (RfDAA) 的设计、合成和应用，能够实时、连续地跟踪转肽反应。这些探针可以通过可视化活细胞中的转肽酶反应以及体外 D,D-和 L,D-转肽酶和分选酶的实时活性测定来实时监测肽聚糖的生物合成。

更新日期：2019-02-26

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