Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, United States.
Department of Chemistry, Boston College, Chestnut Hill, Massachusetts 02467, United States.
ACS Chem Biol. 2023 Sep 15;18(9):1909-1914. doi: 10.1021/acschembio.3c00360. Epub 2023 Aug 10.
The natural product holomycin contains a unique cyclic ene-disulfide and exhibits broad-spectrum antimicrobial activities. Reduced holomycin chelates metal ions with a high affinity and disrupts metal homeostasis in the cell. To identify cellular metalloproteins inhibited by holomycin, reactive-cysteine profiling was performed using isotopic tandem orthogonal proteolysis-activity-based protein profiling (isoTOP-ABPP). This chemoproteomic analysis demonstrated that holomycin treatment increases the reactivity of metal-coordinating cysteine residues in several zinc-dependent and iron-sulfur cluster-dependent enzymes, including carbonic anhydrase II and fumarase A. We validated that holomycin inhibits fumarase A activity in bacterial cells and diminishes the presence of iron-sulfur clusters in fumarase A. Whole-proteome abundance analysis revealed that holomycin treatment induces zinc and iron starvation and cellular stress. This study suggests that holomycin inhibits bacterial growth by impairing the functions of multiple metalloenzymes and sets the stage for investigating the impact of metal-binding molecules on metalloproteomes by using chemoproteomics.
天然产物霍洛霉素含有独特的环烯二硫醚，具有广谱的抗菌活性。还原的霍洛霉素与金属离子高亲和力螯合，并破坏细胞内的金属稳态。为了鉴定被霍洛霉素抑制的细胞金属蛋白酶，使用同位素串联正交蛋白酶解-基于活性的蛋白质谱（isoTOP-ABPP）进行了反应性半胱氨酸分析。这种化学蛋白质组学分析表明，霍洛霉素处理增加了几种锌依赖和铁硫簇依赖酶中金属配位半胱氨酸残基的反应性，包括碳酸酐酶 II 和延胡索酸酶 A。我们验证了霍洛霉素在细菌细胞中抑制延胡索酸酶 A 的活性，并减少延胡索酸酶 A 中铁硫簇的存在。全蛋白质组丰度分析表明，霍洛霉素处理诱导锌和铁饥饿和细胞应激。这项研究表明，霍洛霉素通过损害多种金属酶的功能来抑制细菌生长，并为通过化学蛋白质组学研究金属结合分子对金属蛋白酶组的影响奠定了基础。
