Repurposing Acitretin as an Antipseudomonal Agent Targeting the Pseudomonas aeruginosa Iron-Regulated Heme Oxygenase.
Repurposing Acitretin as an Antipseudomonal Agent Targeting the Pseudomonas aeruginosa Iron-Regulated Heme Oxygenase.
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DOI：
10.1021/acs.biochem.0c00895
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发表时间：
2021-02
期刊：
影响因子：
2.9
通讯作者：
Elizabeth Robinson;A. Wilks;F. Xue
中科院分区：
文献类型：
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作者：
Elizabeth Robinson;A. Wilks;F. Xue
铁是细菌病原体铜绿假单胞菌的生存和毒力所必需的微量营养素。为了克服铁抑制并成功地定殖宿主，铜绿假单胞菌使用多种机制来获得铁，包括分泌高亲和力铁螯合剂（铁载体）或摄取和利用血红素。铜绿假单胞菌血红素加氧酶（HemO）在血红素的感知、摄取和利用中起关键作用，并且已经成为开发抗假单胞菌剂的治疗靶点。使用高通量荧光猝灭试验结合最小抑制浓度测量，我们筛选了Selleck生物活性收集的2100种化合物，并确定了阿维A，一种食品和药物管理局批准的口服类维生素A，作为一种有效的和选择性的HemO抑制剂。阿维A与HemO结合的KD值为0.10 ± 0.02 μM，抑制铜绿假单胞菌PAO 1生长的IC 50为70 ± 18 μg/mL。此外，阿维A对HemO显示出良好的选择性，HemO独特地产生BVIXβ/δ，优于人血红素加氧酶（hHO 1）和其他产生BVIXα的同系物，如脑膜炎奈瑟菌（nmHO）和鲍曼不动杆菌（abHO）的血红素加氧酶。通过1H-15 N杂原子单量子相干核磁共振证实了阿维A在HemO活性位点内的结合，并且分子建模提供了对阿维A与β/δ选择性HemO中对定向血红素具有特异性的残基的潜在相互作用的进一步了解。此外，在20 μM时，阿维A抑制铜绿假单胞菌细胞中HemO的酶活性>60%，并有效阻断铜绿假单胞菌感知和获得血红素的能力，如β-半乳糖苷酶转录报告基因测定所示。
Iron is an essential micronutrient for the survival and virulence of the bacterial pathogen Pseudomonas aeruginosa. To overcome iron withholding and successfully colonize a host, P. aeruginosa uses a variety of mechanisms to acquire iron, including the secretion of high-affinity iron chelators (siderophores) or the uptake and utilization of heme. P. aeruginosa heme oxygenase (HemO) plays pivotal roles in heme sensing, uptake, and utilization and has emerged as a therapeutic target for the development of antipseudomonal agents. Using a high-throughput fluorescence quenching assay combined with minimum inhibitory concentration measurements, we screened the Selleck Bioactive collection of 2100 compounds and identified acitretin, a Food and Drug Administration-approved oral retinoid, as a potent and selective inhibitor of HemO. Acitretin binds to HemO with a KD value of 0.10 ± 0.02 μM and inhibits the growth of P. aeruginosa PAO1 with an IC50 of 70 ± 18 μg/mL. In addition, acitretin showed good selectivity for HemO, which uniquely generates BVIXβ/δ, over human heme oxygenase (hHO1) and other BVIXα-producing homologues such as the heme oxygenases from Neisseria meningitidis (nmHO) and Acinetobacter baumannii (abHO). The binding of acitretin within the HemO active site was confirmed by 1H-15N heteronuclear single-quantum coherence nuclear magnetic resonance, and molecular modeling provided further insight into potential interactions of acitretin with residues specific for orienting heme in the β/δ selective HemO. Moreover, at 20 μM, acitretin inhibited the enzymatic activity of HemO in P. aeruginosa cells by >60% and effectively blocked the ability of P. aeruginosa to sense and acquire heme as demonstrated in the β-galactosidase transcriptional reporter assay.