Fig. 8: P17 tag increased thermostability of G12-scFv. | Nature Communications

Fig. 8: P17 tag increased thermostability of G12-scFv.

From: A 33-residue peptide tag increases solubility and stability of Escherichia coli produced single-chain antibody fragments

Fig. 8

a Reduction of intradomain disulfide bonds of G12-scFv-HA protein by DTT decreased melting temperature (Tm). Five μM G12-scFv-HA protein, treated with 5 mM DTT for 30 min at room temperature or not, was mixed with 1× SYPRO Orange. The fluorescence signal emitted at 570 nm by SYPRO Orange at each temperature was recorded by a QuantStudio PCR apparatus and normalized to the maximal signal reached which was set at 100%. Tm of scFv was determined by three independent thermal shift assays as described in “Methods”. b Tm of G12-scFv-HA and G12-scFv-HA-P17 proteins. c Tm of G12-scFv-HA protein at the presence of in vitro synthesized P17 peptide. After separately mixing with 5 μM, 25 μM, and 50 μM P17 peptide, Tm of 5 μM of G12-scFv-HA protein was measured three times by thermal shift assay. Results shown by average fluorescence values and error bars at each temperature were plotted using Origin 8.0 software.

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