Optimization of Fermentation Process for Production of Elastin-Like and Squid Ring Teeth Fusion Proteins for the Construction of Robust Fibers in Escherichia coli
The recombinant structural protein SRT-ELP36 consisted of repetitive squid ring teeth protein segment and cationic elastin-like polypeptide sequence(PAATAVSHTTHHAP-VPGVG(VPGKG)5)can be used to fabricate protein fibers with extraordinary mechanical properties and biocompatibility.In order to improve the production of recombinant protein SRT-ELP36,the pET-25b(+)vector carrying SRT-ELP36 expression cassette was transformed into Escherichia coli,and the fermentation process was optimized in shake flasks and bioreactors.In comparison with the recombinant E.coli BLR(DE3),BL21(DE3)grew more rapidly with a higher expression level of SRT-ELP36.Thus,batch cultivation of recombinant BL21(DE3)in shake flasks was optimized to achieve the highest cell density OD600 of 22.3 and SRT-ELP36 production of 0.60 g/L in TB(Terrific Broth)medium with 5％volume ratio of medium to flask after 0.5 mmol/L IPTG induction at 37℃ at the end of exponential growth phase.Subsequently,the fed-batch fermentation process in a 5 L bioreactor was developed:induction with 0.5 mmol/L IPTG at the OD600 of 35,a high OUR level up to(180±5)mmol/(L·h).With the highest OD600 of 88,the highest volumetric and specific production of SRT-ELP36 of 1.85 g/L and 70 mg/g was obtained,respectively.Based on the cultivation conditions optimized in the 5 L bioreactor,high cell density fermentation of recombinant BL21(DE3)was successfully carried out in a 50 L bioreactor with the highest OD600 of 103.SRT-ELP36 volumetric and specific production were increased to 2.22 g/L and 90 mg/g,respectively,which was the highest expression level of Squid ring teeth protein as reported to date and providing the foundation for eventual application of SRT-ELP36 in industrial fabrication of mechanically strong fibers.
Escherichia colisquid ring teeth proteinelastin-like polypeptidestructural proteinfermentation optimization
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