Squalene,a natural triterpenoid widely used in cosmetics and pharmaceuticals,can be efficiently biosynthesized using acetyl coenzyme A(acetyl-CoA)and NADPH,which are abundant in Schizochytrium sp.Our previous studies ...Squalene,a natural triterpenoid widely used in cosmetics and pharmaceuticals,can be efficiently biosynthesized using acetyl coenzyme A(acetyl-CoA)and NADPH,which are abundant in Schizochytrium sp.Our previous studies have shown that iron deficiency enhanced squalene production but inhibited growth in Schizochytrium sp.Although the modest reactive oxygen species levels generated under such stress conditions can activate antioxidant defenses and stimulate the synthesis of terpenoids,we propose that the compensatory capacity of the antioxidant system under prolonged iron deficiency may eventually lead to a metabolic imbalance,resulting in the accumulation of fragmentation products.To address this challenge,sesamol was identified as the optimal antioxidant,achieving 199.98 mg/g lipid squalene at 1.5 mmol/L.Transcriptomics revealed that sesamol increased NADPH supply and upregulated ABC-G protein expression to enhance transmembrane transport efficiency.Rational validation through ABC-G2 protein overexpression combined with the in-situ extraction further improved squalene yield to 294.45 mg/g lipid.Furthermore,promoter engineering of ferric-chelate reductase enhanced squalene yield to 354.81 mg/g lipid.Finally,a fermentation strategy combining a single addition of 2 mmol/L sesamol at the start of fermentation with a continuous supplementation of 3 mmol/L sesamol at the logarithmic period of fermentation in a 5 L bioreactor achieved a record-breaking squalene yield of 489.57 mg/g lipid,representing the highest squalene yield reported for Schizochytrium sp.to date.This study provides innovative genetic and bioprocess strategies for industrial-scale squalene production.展开更多
基金supported by the National Key Research and Development Program of China,China(2023YFA0914400)the National Natural Science Foundation of China,China(32422068)+1 种基金the Natural Science Foundation of Jiangsu Province,China(BK20230059)the National Natural Science Foundation of China,China(22378209 and 22038007).
文摘Squalene,a natural triterpenoid widely used in cosmetics and pharmaceuticals,can be efficiently biosynthesized using acetyl coenzyme A(acetyl-CoA)and NADPH,which are abundant in Schizochytrium sp.Our previous studies have shown that iron deficiency enhanced squalene production but inhibited growth in Schizochytrium sp.Although the modest reactive oxygen species levels generated under such stress conditions can activate antioxidant defenses and stimulate the synthesis of terpenoids,we propose that the compensatory capacity of the antioxidant system under prolonged iron deficiency may eventually lead to a metabolic imbalance,resulting in the accumulation of fragmentation products.To address this challenge,sesamol was identified as the optimal antioxidant,achieving 199.98 mg/g lipid squalene at 1.5 mmol/L.Transcriptomics revealed that sesamol increased NADPH supply and upregulated ABC-G protein expression to enhance transmembrane transport efficiency.Rational validation through ABC-G2 protein overexpression combined with the in-situ extraction further improved squalene yield to 294.45 mg/g lipid.Furthermore,promoter engineering of ferric-chelate reductase enhanced squalene yield to 354.81 mg/g lipid.Finally,a fermentation strategy combining a single addition of 2 mmol/L sesamol at the start of fermentation with a continuous supplementation of 3 mmol/L sesamol at the logarithmic period of fermentation in a 5 L bioreactor achieved a record-breaking squalene yield of 489.57 mg/g lipid,representing the highest squalene yield reported for Schizochytrium sp.to date.This study provides innovative genetic and bioprocess strategies for industrial-scale squalene production.
