Abstract
Poly-γ-glutamic acid (γ-PGA) and nattokinase (NK) are the main substances produced by Bacillus subtilis natto in solid-state fermentation and have wide application prospects. We found that our strains had higher activity of nattokinase when soybeans were used as substrate to increase the yield of γ-PGA. Commercial production of γ-PGA and nattokinase requires an understanding of the mechanism of co-production. Here, we obtained the maximum γ-PGA yield (358.5 g/kg, w/w) and highest activity of NK during fermentation and analyzed the transcriptome of Bacillus subtilis natto during co-production of γ-PGA and NK. By comparing changes in expression of genes encoding key enzymes and the metabolic pathways associated with the products in genetic engineering, the mechanism of co-production of γ-PGA and nattokinase can be summarized based on RNA-seq analysis. This study firstly provides new insights into the mechanism of co-production of γ-PGA and nattokinase by Bacillus subtilis natto and reveals potential molecular targets to promote the co-production of γ-PGA and nattokinase.
Highlights
Poly-γ-glutamic acid is a natural multifunctional biopolymer composed of D- and/or L-glutamic acid units linked by γ-amide bonds [1]
The activity of nattokinase remained stable constant after 24 h, but the concentration of γ-PGA decreased significantly after 36 h. This may be caused by γ-PGA depolymerase, which is heavily excreted by B. subtilis natto in the late stationary phase [40]
By analyzing the change of γ-PGA concentration and nattokinase activity at different periods, it was found that the synthesis and accumulation of γ-PGA occurred mainly in the logarithmic growth phase, which is consistent with the study of Mahaboob Ali [41]
Summary
Poly-γ-glutamic acid is a natural multifunctional biopolymer composed of D- and/or L-glutamic acid units linked by γ-amide bonds [1]. Nattokinase is a biological enzyme extracted from fermented soybeans [9]. It is a linear chain of amino acid chains with spatial folding, produced in the form of a signal peptide, a propeptide and a mature peptide, with a molecular weight of 27.7 KDa [10]. The fibrinolytic activity of NK is 4 times higher than that of plasmin and its molecular weight is much smaller than that of urokinase and lumbrokinase. It is well absorbed by the intestine [12,13,14]. Nattokinase is currently considered as a potential drug for the preventing and treatment of cardiovascular diseases [15, 16]
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