New Insight into the Mechanism of Anaerobic Heme Degradation
New Insight into the Mechanism of Anaerobic Heme Degradation
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DOI：
10.1021/acs.biochem.9b00841
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发表时间：
2019-11-19
期刊：
影响因子：
2.9
通讯作者：
Lanzilotta, William N.
中科院分区：
文献类型：
--
作者：
Mathew, Liju G.;Beattie, Nathaniel R.;Lanzilotta, William N.
ChuW、ChuX和ChuY是一个启动子下游的连续基因，当铁限制时，它们在肠道病原体大肠杆菌O157:H7中表达。这些基因和相应的蛋白质是血红素摄取和利用操纵子的一部分，这是其他几种肠道病原体（如霍乱弧菌）所共有的。血红素的有氧降解已经在人类和几种致病菌（包括大肠杆菌O157:H7）中得到了很好的表征，但直到最近才发现ChuW催化血红素的厌氧降解以释放铁并产生活性四吡啶，称为“厌氧素”。ChuY已被证明具有厌氧素还原酶的功能，其作用与胆绿素还原酶相似。在这项工作中，我们采用了生化和生物物理的方法来进一步探讨血红素厌氧降解的机制。我们证明血红素的铁原子不参与ChuW的催化机制，s -腺苷- l-蛋氨酸结合诱导有利于催化的构象变化。此外，我们发现ChuX和ChuY对ChuW的流失率具有协同效应和加性效应。最后，我们发现在厌氧条件下，ChuS是ChuW血红素或原卟啉IX的有效来源。这些数据表明，ChuS在体内可能具有双重功能。具体来说，ChuS在血红素的有氧代谢过程中充当血红素加氧酶，但在厌氧条件下充当细胞质血红素储存蛋白，类似于铜绿假单胞菌的PhuS（45%序列同一性）。
ChuW, ChuX, and ChuY are contiguous genes downstream from a single promoter that are expressed in the enteric pathogen Escherichia coli O157:H7 when iron is limiting. These genes, and the corresponding proteins, are part of a larger heme uptake and utilization operon that is common to several other enteric pathogens, such as Vibrio cholerae. The aerobic degradation of heme has been well characterized in humans and several pathogenic bacteria, including E. coli O157:H7, but only recently was it shown that ChuW catalyzes the anaerobic degradation of heme to release iron and produce a reactive tetrapyrrole termed "anaerobilin". ChuY has been shown to function as an anaerobilin reductase, in a role that parallels biliverdin reductase. In this work we have employed biochemical and biophysical approaches to further interrogate the mechanism of the anaerobic degradation of heme. We demonstrate that the iron atom of the heme does not participate in the catalytic mechanism of ChuW and that S-adenosyl-L- methionine binding induces conformational changes that favor catalysis. In addition, we show that ChuX and ChuY have synergistic and additive effects on the turnover rate of ChuW. Finally, we have found that ChuS is an effective source of heme or protoporphyrin IX for ChuW under anaerobic conditions. These data indicate that ChuS may have dual functionality in vivo. Specifically, ChuS serves as a heme oxygenase during aerobic metabolism of heme but functions as a cytoplasmic heme storage protein under anaerobic conditions, akin to what has been shown for PhuS (45% sequence identity) from Pseudomonas aeruginosa.