Expression, Purification, and In Silico Characterization of Mycobacterium smegmatis Alternative Sigma Factor SigB

Figure 3

Temperature optimization for the expression of recombinant His-SigB protein. (a) 10% SDS PAGE depicting recombinant His-SigB protein expression profile in CodonPlus (DE3) cells at temperatures: 37°C, 25°C, and 16°C with 0.1 mM IPTG. The highest protein expression was observed at 37°C. (b) 10% SDS PAGE depicting distribution of recombinant His-SigB protein in cell pellet and supernatant fractions at different temperatures. The highest protein solubility was observed at 25°C supernatant fraction. M: marker; V.O.: pET28a vector only control; U: uninduced His-SigB clone; P: pellet; S: supernatant. (c) Gel-based semiquantitative analysis from (a) depicts the highest expression of His-SigB at 37°C with 0.1 mM IPTG concentration. (d) Semiquantitative analysis from (b) depicts higher relative protein solubility observed in the supernatant fraction at 25°C with 0.1 mM IPTG. Semiquantitative analysis of the gels was performed with Quantity One software (BIO-RAD).

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