Mycobacterium tuberculosis WhiB3 responds to O2 and nitric oxide via its [4Fe-4S] cluster and is essential for nutrient starvation survival
Mycobacterium tuberculosis WhiB3 responds to O2 and nitric oxide via its [4Fe-4S] cluster and is essential for nutrient starvation survival
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DOI：
10.1073/pnas.0700490104
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发表时间：
2007-07-10
影响因子：
11.1
通讯作者：
Steyn, Adrie J. C.
中科院分区：
文献类型：
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作者：
Singh, Amit;Guidry, Loni;Steyn, Adrie J. C.
结核分枝杆菌（Mtb）氧化还原生物学的一个基本挑战是了解参与感知氧化还原信号（如氧（O-2），一氧化氮（NO）和营养耗尽）的机制，这些信号被认为在持久性中起着至关重要的作用。在这里，我们表明，Mtb WhiB 3响应休眠信号NO和O-2通过其铁硫（Fe-S）簇。为了在功能上组装WhiB 3 Fe-S簇，我们鉴定并表征了Mtb半胱氨酸脱硫酶（IscS; Rv 3025 c），并开发了用于在Mtb中组装Fe-S簇的天然酶促重建系统。还原的WhiB 3的EPR和UV-可见光谱分析与EPR沉默[4Fe-4S]（2+）到EPR可见[4Fe-4S]（+）的单电子还原一致。大气中的O-2逐渐降解WhiB 3 [4Fe-4S]2（+）簇合物，生成[3Fe-4S]（+）中间体。此外，EPR分析表明，NO与Fe-S簇形成蛋白质结合的二亚硝基铁二硫醇复合物，表明NO特异性靶向WhiB 3 Fe-S簇。我们的数据表明，WhiB 3 4Fe-4S团簇的降解机制类似于富马酸盐硝酸盐调节剂。重要的是，Mtb Delta whiB 3在乙酸盐培养基上显示出增强的生长，但在含有葡萄糖、丙酮酸盐、琥珀酸盐或富马酸盐作为唯一碳源的培养基上显示出生长缺陷。我们的研究结果牵连WhiB 3在代谢开关和在传感生理相关的主机信号分子NO和O-2通过其[4Fe-4S]簇。两者合计，我们的研究结果表明，WhiB 3是一个细胞内的氧化还原传感器，整合环境氧化还原信号与核心中间代谢。
A fundamental challenge in the redox biology of Mycobacterium tuberculosis (Mtb) is to understand the mechanisms involved in sensing redox signals such as oxygen (O-2), nitric oxide (NO), and nutrient depletion, which are thought to play a crucial role in persistence. Here we show that Mtb WhiB3 responds to the dormancy signals NO and O-2 through its iron-sulfur (Fe-S) cluster. To functionally assemble the WhiB3 Fe-S cluster, we identified and characterized the Mtb cysteine desulfurase (IscS; Rv3025c) and developed a native enzymatic reconstitution system for assembling Fe-S clusters in Mtb. EPR and UV-visible spectroscopy analysis of reduced WhiB3 is consistent with a one-electron reduction of EPR silent [4Fe-4S](2+) to EPR visible [4Fe-4S](+). Atmospheric O-2 gradually degrades the WhiB3 [4Fe-4S]2(+) cluster to generate a [3Fe-4S](+) intermediate. Furthermore, EPR analysis demonstrates that NO forms a protein-bound dinitrosyl-iron-dithiol complex with the Fe-S cluster, indicating that NO specifically targets the WhiB3 Fe-S cluster. Our data suggest that the mechanism of WhiB3 4Fe-4S cluster degradation is similar to that of fumarate nitrate regulator. Importantly, Mtb Delta whiB3 shows enhanced growth on acetate medium, but a growth defect on media containing glucose, pyruvate, succinate, or fumarate as the sole carbon source. Our results implicate WhiB3 in metabolic switching and in sensing the physiologically relevant host signaling molecules NO and O-2 through its [4Fe-4S] cluster. Taken together, our results suggest that WhiB3 is an intracellular redox sensor that integrates environmental redox signals with core intermediary metabolism.