State Key Laboratory of Elemento-Organic Chemistry and Department of Chemical Biology, National Pesticide Engineering Research Center (Tianjin), Nankai University, Tianjin, China.
Tianjin Key Laboratory of Technologies Enabling Development of Clinical Therapeutics and Diagnostics (Theranostics), School of Pharmacy, Tianjin Medical University, Tianjin, China.
Proteins. 2023 Aug;91(8):1163-1172. doi: 10.1002/prot.26501. Epub 2023 Apr 27.
Coproporphyrinogen oxidase (CPO) plays important role in the biosynthesis of heme by catalyzing the coproporphyrinogen III to coproporphyrin III. However, in earlier research, it was regarded as the protoporphyrinogen oxidase (PPO) because it can also catalyze the oxidation of protoporphyrinogen IX to protoporphyrin IX. Identification of the commonalities in CPO and PPO would help us to get a further understanding of the enzyme function. In this work, we explored the role of a non-conserved residue, Asp65 in Bacillus subtilis CPO (bsCPO), whose corresponding residues in PPO from various species are neutral or positive residue (arginine in human PPO or asparagine in tobacco PPO, etc.). We found that Asp65 performs its function by forming a polar interaction network with its surrounding residues in bsCPO, which is important for enzymatic activity. This polar network maintains the substrate binding chamber and stabilizes the micro-environment of the isoalloxazine ring of FAD for the substrate-FAD interaction. Both the comparison of the crystal structures of bsCPO with PPO and our previous work showed that a similar polar interaction network is also present in PPOs. The results confirmed our conjecture that non-conserved residues can form a conserved element to maintain the function of CPO or PPO.
粪卟啉原氧化酶（CPO）在血红素的生物合成中发挥重要作用，通过催化粪卟啉原 III 生成粪卟啉 III。然而，在早期的研究中，由于它还可以催化原卟啉原 IX 的氧化生成原卟啉 IX，因此它被认为是原卟啉原氧化酶（PPO）。确定 CPO 和 PPO 之间的共同特征将有助于我们进一步了解酶的功能。在这项工作中，我们探索了一个非保守残基 Asp65 在枯草芽孢杆菌 CPO（bsCPO）中的作用，该残基在来自不同物种的 PPO 中的对应残基是中性或正电荷残基（人 PPO 中的精氨酸或烟草 PPO 中的天冬酰胺等）。我们发现 Asp65 通过与 bsCPO 周围的残基形成极性相互作用网络来发挥其功能，这对于酶活性很重要。该极性网络维持了底物结合腔，并稳定了 FAD 的异咯嗪环的微环境，有利于底物-FAD 相互作用。bsCPO 与 PPO 的晶体结构比较以及我们之前的工作表明，类似的极性相互作用网络也存在于 PPO 中。结果证实了我们的假设，即非保守残基可以形成一个保守元素来维持 CPO 或 PPO 的功能。
