Southam Hannah M, Williamson Michael P, Chapman Jonathan A, Lyon Rhiannon L, Trevitt Clare R, Henderson Peter J F, Poole Robert K
Department of Molecular Biology and Biotechnology, The University of Sheffield, Western Bank, Sheffield S10 2TN, UK.
Centre for Bacterial Cell Biology, Medical School, Newcastle University, Newcastle upon Tyne NE2 4AX, UK.
Antioxidants (Basel). 2021 Jun 5;10(6):915. doi: 10.3390/antiox10060915.
Carbon monoxide (CO)-releasing molecules (CORMs) are used to deliver CO, a biological 'gasotransmitter', in biological chemistry and biomedicine. CORMs kill bacteria in culture and in animal models, but are reportedly benign towards mammalian cells. CORM-2 (tricarbonyldichlororuthenium(II) dimer, RuCl(CO)), the first widely used and commercially available CORM, displays numerous pharmacological, biochemical and microbiological activities, generally attributed to CO release. Here, we investigate the basis of its potent antibacterial activity against and demonstrate, using three globin CO sensors, that CORM-2 releases negligible CO (<0.1 mol CO per mol CORM-2). A strong negative correlation between viability and cellular ruthenium accumulation implies that ruthenium toxicity underlies biocidal activity. Exogenous amino acids and thiols (especially cysteine, glutathione and N-acetyl cysteine) protected bacteria against inhibition of growth by CORM-2. Bacteria treated with 30 μM CORM-2, with added cysteine and histidine, exhibited no significant loss of viability, but were killed in the absence of these amino acids. Their prevention of toxicity correlates with their CORM-2-binding affinities (Cys, 3 μM; His, 130 μM) as determined by H-NMR. Glutathione is proposed to be an important intracellular target of CORM-2, with CORM-2 having a much higher affinity for reduced glutathione (GSH) than oxidised glutathione (GSSG) (GSH, 2 μM; GSSG, 25,000 μM). The toxicity of low, but potent, levels (15 μM) of CORM-2 was accompanied by cell lysis, as judged by the release of cytoplasmic ATP pools. The biological effects of CORM-2 and related CORMs, and the design of biological experiments, must be re-examined in the light of these data.
一氧化碳释放分子（CORMs）在生物化学和生物医学中用于递送生物“气体递质”一氧化碳（CO）。CORMs在培养物和动物模型中可杀死细菌，但据报道对哺乳动物细胞无害。CORM-2（二氯三羰基钌（II）二聚体，RuCl(CO)）是首个被广泛使用且可商购的CORM，具有多种药理、生化和微生物学活性，这些活性通常归因于CO的释放。在此，我们研究了其对[具体细菌名称未给出]的强效抗菌活性的基础，并使用三种球蛋白CO传感器证明，CORM-2释放的CO可忽略不计（每摩尔CORM-2释放<0.1摩尔CO）。活力与细胞钌积累之间存在强烈的负相关，这表明钌毒性是杀菌活性的基础。外源性氨基酸和硫醇（尤其是半胱氨酸、谷胱甘肽和N-乙酰半胱氨酸）可保护细菌免受CORM-2对生长的抑制。用30μM CORM-2处理并添加半胱氨酸和组氨酸的细菌，其活力没有显著损失，但在没有这些氨基酸的情况下会被杀死。它们对毒性的预防与其通过H-NMR测定的CORM-2结合亲和力相关（半胱氨酸，3μM；组氨酸，130μM）。谷胱甘肽被认为是CORM-2的重要细胞内靶点，CORM-2对还原型谷胱甘肽（GSH）的亲和力远高于氧化型谷胱甘肽（GSSG）（GSH，2μM；GSSG，25,000μM）。低浓度但强效（15μM）的CORM-2的毒性伴随着细胞裂解，这可通过细胞质ATP池的释放来判断。鉴于这些数据，必须重新审视CORM-2及相关CORMs的生物学效应以及生物学实验的设计。
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