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Fig. 4 | Microbial Cell Factories

Fig. 4

From: Evaluation of factors influencing expression and extraction of recombinant bacteriophage endolysins in Escherichia coli

Fig. 4

Solubility N-term tags performance on soluble endolysin yields. A, B SDS PAGE showing the position of each fusion tag to recombinant endolysins; C, D E. coli C43(DE3) harboring pET-32a modified with N-term solubility tags (DsbC, Sumo and Trx) in frame fusions to MatN/Cg genes was induced at 20 °C by addition of 1 mM IPTG. Cells were harvested after 20 h of incubation with constant shaking and processed to obtain soluble (S) and insoluble (I) fractions. Equivalent protein amounts (10 µg) of each sample were loaded on 12% SDS-PAGE. Upon gel electrophoresis, gels were stained with Coomassie Brilliant Blue R-250 and unstained following standard protocols. Control (CTL) lanes correspond to cellular lysates of uninduced E. coli C43(DE3) harboring pET32a-MatN/Cg. E, F Total amounts of recombinant endolysin (column height) and the ratio of soluble/insoluble protein (SP, gray dotted pattern; IP, diagonal lines pattern) were calculated after background normalization with ImageJ, open-source image processing software. Endolysin expression and solubility were analyzed in biological triplicates. Error bars represent SD of mean values. **p  <  0.005, *p  <  0.05 (total endolysin expression of N-His group vs. N-Tags groups)

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