Co-expression of recombinant human epidermal growth factor (rhEGF) in Escherichia coli BL21 (DE3) with Bacillus cereus phospholipase C

Indriyani, Annisa; Gaffar, Shabarni; Latifah, F. P. U.; Maksum, IP

doi:10.1063/5.0002067

Cited by 1 publication

(1 citation statement)

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“…Most of prethrombin-2 (PT-2) produced in E. coli has minor activity, due to formation of inclusion bodies (Soejima et al, 2001). Therefore, simple strategies to overcome inclusion bodies formation were needed, such as: 1) Intracellular expression with refolding of fusion protein using E. coli Artic express (Maksum et al, 2019;S Silaban et al, 2019a;Silaban et al, 2019b) or coexpression using chaperone (Gupta, 2002;Silaban et al, 2014;Rizkia et al, 2015;Silaban et al, 2019c), 2) Extracellular expression using signal peptide and phospholipase co-expression (Sriwidodo et al, 2017;Maksum et al, 2017c;Melati et al, 2019;Indriyani et al, 2019;Indriyani et al, 2020;Latifah et al, 2020) .…”

Section: Introductionmentioning

confidence: 99%

The Effect of Single Co-expression of The DnaK-DnaJ-GrpE and GroEL/ES Chaperones and Their Combination on Expression Intein-pretrombin-2 in Escherichia coli ER2566

Maksum

Wildan

Hasan

et al. 2020

Valensi

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The use of recombinant thrombin in the manufacture of fibrin glue allows diseases contamination to be avoided. However, the expression of recombinant protein in E. coli still has a disadvantage of the formation of inclusion bodies, so it needs to be minimized by co-expression of chaperones. Therefore, the aim of this study was to determine the effect of single DnaK-DnaJ-GrpE and GroEL/ES chaperone expression and their combination on the expression of intein-pretrombin-2Ti,pH on E. coli ER2566. The method started with isolation of pTWIN1-prethrombin-2Ti,pH and pG-KJE8 from E. coli TOP10F' and DH5α respectively, the co-transformation of the expression host E. coli ER2566 using pG-KJE8 and pTWIN1-prethrombin-2Ti,pHvectors, the chaperone co-expression was induced using L-Arabinosa before IPTG induction and cell culture growth was incubated at 22 oC. The expression products were characterized by using Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The results of the co-expression of chaperone showed that the number of soluble fraction was higher than the one without co-expression of chaperone. In addition, the co-expression of chaperone using pG-KJE8 in intein-prothrombin-2Ti,pH expression was sufficient using tetracycline as an inducer.

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