Motz Rachel N, Anderson Jaden K, Nolan Elizabeth M
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, United States.
Biochemistry. 2025 Jan 7;64(1):224-237. doi: 10.1021/acs.biochem.4c00581. Epub 2024 Dec 24.
The pathogen-associated -glucosyltransferase IroB is involved in the biosynthesis of salmochelins, -glucosylated derivatives of enterobactin (Ent), which is a triscatecholate siderophore of enteric bacteria including and . Here, we reassess the ability of IroB to -glucosylate non-native triscatecholate mimics of Ent, which may have utility in the design and development of siderophore-based therapeutics and diagnostics. We establish TRENCAM (TC) and MECAM (MC), synthetic Ent analogs with tris(2-aminoethyl)amine- or mesitylene-derived backbones replacing the trilactone core of Ent, respectively, and their monoglucosylated congeners as substrates of IroB. Time course analyses and steady-state kinetic studies, which were performed under conditions that provide enhanced activity relative to prior studies, inform the substrate selectivity and catalytic efficiencies of this enzyme. We extend these findings to the preparation of a siderophore-antibiotic conjugate composed of monoglucosylated TC and ampicillin (MGT-Amp). Examination of its antibacterial activity and receptor specificity demonstrates that MGT-Amp targets pathogenicity because it shows specificty for the pathogen-associated outer membrane receptor IroN. Overall, our findings extend the biochemical characterization of IroB and its substrate scope and illustrate the ability to leverage a bacterial -glucosyltransferase for non-native chemoenzymatic transformations along with potential applications of salmochelin mimics.
病原体相关的β-葡萄糖基转移酶IroB参与了沙门菌素的生物合成,沙门菌素是肠杆菌素(Ent)的β-葡萄糖基化衍生物,肠杆菌素是包括大肠杆菌和沙门氏菌在内的肠道细菌的三儿茶酚铁载体。在此,我们重新评估IroB对Ent的非天然三儿茶酚类似物进行β-葡萄糖基化的能力,这可能在基于铁载体的治疗和诊断的设计与开发中具有实用性。我们分别建立了TRENCAM(TC)和MECAM(MC),这两种合成的Ent类似物,其具有分别用三(2-氨基乙基)胺或均三甲苯衍生的主链取代Ent的三内酯核心,以及它们的单葡萄糖基化同系物作为IroB的底物。在相对于先前研究能提高活性的条件下进行的时间进程分析和稳态动力学研究,为该酶的底物选择性和催化效率提供了信息。我们将这些发现扩展到了由单葡萄糖基化的TC和氨苄青霉素(MGT-Amp)组成的铁载体-抗生素缀合物的制备。对其抗菌活性和受体特异性的研究表明MGT-Amp靶向致病性,因为它对病原体相关的外膜受体IroN具有特异性。总体而言,我们的发现扩展了IroB的生化特性及其底物范围,并说明了利用细菌β-葡萄糖基转移酶进行非天然化学酶转化的能力以及沙门菌素类似物的潜在应用。