A Family of Helminth Molecules that Modulate Innate Cell Responses via Molecular Mimicry of Host Antimicrobial Peptides
Figure 4
Expression and CD spectroscopy of recombinant FhHDM-1.
(A) The full-length FhHDM-1 cDNA, minus the N-terminal signal peptide, was expressed in E. coli and the His-tagged recombinant was purified from cell lysates using Ni-NTA agarose (Qiagen). P, pre-column; FT, flow-through; W, wash, E1, imidazole eluate. Co-eluting proteins were removed by RP-HLPC resulting in recombinant FhHDM-1 of very high purity (E2). (B) CD spectra of recombinant 0.1 mg/mL−1 FhHDM-1 at pH 7.3. The wavelength scan was performed between 190 and 250 nm. The final spectrum (closed circles in the absence of 30% (v/v) TFE and open circles in the presence of 30% (v/v) TFE) is the average result from three scans measured at 20°C. The CONTINLL algorithm from the CDPro software package [27] produced the best fit (solid lines) against the SP29 protein database [28] with r.m.s.d. values for all samples ≤0.325. FhHDM-1 adopts a near identical solution structure in both native and recombinant form at both pH 4.5 and pH 7.3 (data not shown). The resulting secondary structure proportions are reported in Table S1.
