Tuning of acyl-ACP thioesterase activity directed for tailored fatty acid synthesis
Tuning of acyl-ACP thioesterase activity directed for tailored fatty acid synthesis
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调节酰基-ACP硫酯酶活性以定制脂肪酸合成
DOI：
10.1007/s00253-018-8770-6
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发表时间：
2018-04-01
影响因子：
5
通讯作者：
Xue, Song
中科院分区：
文献类型：
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作者：
Feng, Yanbin;Zhang, Yunxiu;Xue, Song
近年来，中链脂肪酸作为生物燃料的来源引起了人们的极大关注。酰基-ACP硫酯酶是决定脂肪酸从头合成的关键酶。虽然重组中链酰基-ACP硫酯酶（TE）影响异源细胞中的脂肪酸谱，但仅通过工程化特定TE来定制脂肪酸组合物仍然是棘手的。在这项研究中，C8-C10-特异性硫酯酶FatB 2的活性从细萼距花C10-ACP上的活性是C8-ACP上的两倍高，基于体外合成的C8-C16酰基-ACP池。体内实验表明，ChFatB 2基因工程菌中C8脂肪酸的积累占总脂肪酸的84.9%，而体内实验表明ChFatB 2基因工程菌中C8脂肪酸的积累占总脂肪酸的84.9%。大肠杆菌为了实现C10脂肪酸的生产，ChFatB 2的结构研究和酶分析的基础上合理调整。一个I198 E突变体被确定为重新分配的C8-ACP流，导致C10脂肪酸被生产的主要成分在体内总脂肪酸的57.6%。研究表明，TE相对于β-酮脂酰-ACP合酶（KAS）的活性直接决定了脂肪酸的组成。我们的研究结果提供了一个前瞻性的策略，在定制脂肪酸合成的TE活动的基础上TE-ACP相互作用的调整。
Medium-chain fatty acids have attracted significant attention as sources of biofuels in recent years. Acyl-ACP thioesterase, which is considered as the key enzyme to determine the carbon chain length, catalyzes the termination of de novo fatty acid synthesis. Although recombinant medium-chain acyl-ACP thioesterase (TE) affects the fatty acid profile in heterologous cells, tailoring of the fatty acid composition merely by engineering a specific TE is still intractable. In this study, the activity of a C8–C10-specific thioesterase FatB2 fromCuphea hookerianaon C10-ACP was quantified twice as high as that on C8-ACP based on a synthetic C8–C16 acyl-ACP pool in vitro. Whereas in vivo, it was demonstrated that ChFatB2 preferred to accumulate C8 fatty acids with 84.9% composition in the ChFatB2-engineeredE. colistrain. To achieve C10 fatty acid production, ChFatB2 was rationally tuned based on structural investigation and enzymatic analysis. An I198E mutant was identified to redistribute the C8-ACP flow, resulting in C10 fatty acid being produced as the principal component at 57.6% of total fatty acids in vivo. It was demonstrated that the activity of TE relative to β-ketoacyl-ACP synthases (KAS) directly determined the fatty acid composition. Our results provide a prospective strategy in tailoring fatty acid synthesis by tuning of TE activities based on TE–ACP interaction.