Hemoglobin (Hb) and myoglobin (Mb) have been extensively employed as oxygen carriers, iron supplements, and food additives, but the microbial synthesis of functional Hb and Mb remains a challenge. Herein, we engineered a Corynebacterium glutamicum strain with efficient heme supply for producing active Hb and Mb. Through the iterative multimodule strategy, including improvement of precursor supply (5-aminolevulinate), redirection of unstable hydroxymethylbilane, selection of the coproporphyrin-dependent pathway as the superior downstream route, enhancement of rate-limiting steps, and prevention of heme secretion, the heme titer was increased by 90.75-fold. Combined with the optimized expression of globins, the engineered HEME3-ΔhrtBA strain achieved the highest titers at the shake-flask level for soybean Hb (1.10 g/L), clover Hb (1.12 g/L), bovine Mb (0.56 g/L), and porcine Mb (1.14 g/L), respectively. Notably, the biosynthesized Hb and Mb presented biochemical properties similar to those of their natural counterparts. This engineered strain provides a scalable platform for synthesizing diverse functional hemoproteins.

中文翻译：

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具有有效血红素供应的工程谷氨酸棒状杆菌用于合成高活性血红蛋白和肌红蛋白

血红蛋白 （Hb） 和肌红蛋白 （Mb） 已被广泛用作氧载体、铁补充剂和食品添加剂，但功能性 Hb 和 Mb 的微生物合成仍然是一个挑战。在此，我们设计了一种具有高效血红素供应的谷氨酸棒状杆菌菌株，用于产生活性 Hb 和 Mb。通过迭代多模块策略，包括改善前体供应 （5-氨基乙酰丙酸酯）、重定向不稳定的羟甲基胆烷、选择粪卟啉依赖性途径作为优势下游途径、增强限速步骤和防止血红素分泌，血红素滴度提高了 90.75 倍。结合珠蛋白的优化表达，工程化的 HEME3-ΔhrtBA 菌株在摇瓶水平上分别对大豆 Hb （1.10 g/L）、三叶草 Hb （1.12 g/L）、牛 Mb （0.56 g/L） 和猪 Mb （1.14 g/L） 达到最高滴度。值得注意的是，生物合成的 Hb 和 Mb 表现出与其天然对应物相似的生化性质。这种工程菌株为合成各种功能性血蛋白提供了一个可扩展的平台。