JOURNAL OF DAIRY SCIENCE
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2013
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IF 2.566
Liang Li,Ying Ma
The influence of fatty acids on the β-oxidation system and thioesterase of Lactococcus lactis was investigated in this study. The results showed that fatty acids (C8:0–C16:0) significantly inhibited the growth of Lactococcus lactis , and laurate (C12:0) had the highest bactericidal effects. We detected the maximum activity of β-oxidation at different incubation times (8, 12, and 18 h) to be 6.460, 7.751, and 8.203, respectively, and the maximum activity of thioesterase at different incubation times (8, 12, and 18 h) to be 19.498, 27.180, and 12.800, respectively. Fatty acids were seen to induce the β-oxidation system and activity of thioesterase; decanoic acid (C10:0) and palmitic acid (C16:0) were also seen to induce the β-oxidation system of Lactococcus lactis , but the induced ability was significantly different. Octanoic acid (C8:0) and palmitic acid (C16:0) were seen to induce thioesterase activity in Lactococcus lactis . When 1 m M palmitic acid (C16:0) was added to M17 broth, the activity of thioesterase increased 5-fold after 2 min; however, adding octanoic acid (C8:0) changed the activity little. Evidence showed that the ability to induce the β-oxidation system and thioesterase activity was related to the fatty acids’ chain lengths.
研究领域:
微生物学
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生物化学
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PubMed
高影响因子
JOURNAL OF DAIRY SCIENCE
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2014
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IF 2.55
Liang Li,Ying Ma
The effects of divalent metal ions (Ca 2+ , Mg 2+ , Fe 2+ , and Cu 2+ ) on the growth, β-oxidation system, and thioesterase activity of Lactococcus lactis were investigated. Different metal ions significantly influenced the growth of L. lactis : Ca 2+ and Fe 2+ accelerated growth, whereas Cu 2+ inhibited growth. Furthermore, Mg 2+ inhibited growth of L. lactis at a low concentration but stimulated growth of L. lactis at a high concentration. The divalent metal ions had significant effects on activity of the 4 key enzymes of the β-oxidation system (acyl-CoA dehydrogenase, enoyl-CoA hydratase, L-3-hydroxyacyl-CoA dehydrogenase, and thiolase) and thioesterase of L. lactis. The activity of acyl-CoA dehydrogenases increased markedly in the presence of Ca 2+ and Mg 2+ , whereas it decreased with 1 mmol/L Fe 2+ or 12 mmol/L Mg 2+ . All the metal ions could induce activity of enoyl-CoA hydratase. In addition, 12 mmol/L Mg 2+ significantly stimulated activity of L-3-hydroxyacyl-CoA dehydrogenase, and all metal ions could induce activity of thiolase, although thiolase activity decreased significantly when 0.05 mmol/L Cu 2+ was added into M17 broth. Inhibition of thioesterase activity by all 4 metal ions could be reversed by 2 mmol/L Ca 2+ . These results help us understand the effect of metal ions on the β-oxidation system and thioesterase activity of Lactococcus lactis .
研究领域:
微生物学
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生物化学
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PubMed
高影响因子
METABOLIC ENGINEERING
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2020
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IF 7.263
Xi Deng,Liuqing Chen,Mohan Hei,Tiangang Liu,Yan Feng,Guang-Yu Yang
Medium-chain fatty acids (C6–C10) have attracted much attention recently for their unique properties compared to their long-chain counterparts, including low melting points and relatively higher carbon conversion yield. Thioesterase enzymes , which can catalyze the hydrolysis of acyl-ACP (acyl carrier protein) to release free fatty acids (FAs), regulate both overall FA yields and acyl chain length distributions in bacterial and yeast fermentation cultures. These enzymes typically prefer longer chain substrates. Herein, seeking to increase bacterial production of MCFAs, we conducted structure-guided mutational screening of multiple residues in the substrate-binding pocket of the E. coli thioesterase enzyme ‘TesA. Confirming our hypothesis that enhancing substrate selectivity for medium-chain acyl substrates would promote overall MCFA production, we found that replacement of residues lining the bottom of the pocket with more hydrophobic residues strongly promoted the C8 substrate selectivity of ‘TesA. Specifically, two rounds of saturation mutagenesis led to the identification of the ‘TesA RD−2 variant that exhibited a 133-fold increase in selectivity for the C8-ACP substrate as compared to C16-ACP substrate. Moreover, the recombinant expression of this variant in an E. coli strain with a blocked β -oxidation pathway led to a 1030% increase in the in vivo octanoic acid (C8) production titer. When this strain was fermented in a 5-L fed-batch bioreactor , it produced 2.7 g/L of free C8 (45%, molar fraction) and 7.9 g/L of total free FAs, which is the highest-to-date free C8 titer to date reported using the E. coli type II fatty acid synthetic pathway. Thus, reshaping the substrate binding pocket of a bacterial thioesterase enzyme by manipulating the hydrophobicity of multiple residues altered the substrate selectivity and therefore fatty acid product distributions in cells. Our study demonstrates the relevance of this strategy for increasing titers of industrially attractive MCFAs as fermentation products.
研究领域:
生物化学
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合成生物学
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代谢工程
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高影响因子
MOLECULAR MICROBIOLOGY
·
2021
·Vol. 116
·期 3
·783-793
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IF 3.501
Yidong Zhou,Siyuan Gao,Mingliang Zhang,Wankui Jiang,Zhijian Ke,Jiguo Qiu,Jianhong Xu,Qing Hong
Salicylate is a typical aromatic compound widely distributed in nature. Microbial degradation of salicylate has been well studied and salicylate hydroxylases play essential roles in linking the peripheral and ring-cleavage catabolic pathways. The direct hydroxylation of salicylate catalyzed by salicylate-1-hydroxylase or salicylate-5-hydroxylase has been well studied. However, the CoA mediated salicylate 5-hydroxylation pathway has not been characterized in detail. Here, we elucidate the molecular mechanism of the reaction in the conversion of salicylate to gentisate in the carbaryl-degrading strain Rhizobium sp. X9. Three enzymes (salicylyl-CoA ligase CehG, salicylyl-CoA hydroxylase CehH and gentisyl-CoA thioesterase CehI) catalyzed the conversion of salicylate to gentisate via a route, including CoA thioester formation, hydroxylation and thioester hydrolysis. Further analysis indicated that genes cehGHI are also distributed in other bacteria from terrestrial environment and marine sediments. These genomic evidences highlight the role of this salicylate degradation pathway in the carbon cycle of soil organic compounds and marine sediments. Our findings of this three-step strategy enhanced the current understanding of CoA mediated degradation of salicylate.
研究领域:
微生物学
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环境科学
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生物化学
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PubMed
高影响因子
危险品化学品经营许可证（带存储）