A Family of Helminth Molecules that Modulate Innate Cell Responses via Molecular Mimicry of Host Antimicrobial Peptides
Figure 1
Identification and characterisation of native FhHDM-1.
(A) Secretory proteins collected from adult F. hepatica following in vitro culture were separated by gel filtration and the resulting high molecular mass (>200 kDa) peak (peak I; PI) was separated further using reverse phase HLPC (RP-HPLC). Fractions collected following gel filtration and RP-HPLC were run on reducing 4–12% Bis-Tris gels (B) and showed that a prominent ∼ 6 kDa protein present in total adult secretory proteins (S) was enriched in PI and purified to homogeneity (>95%) following RP-HPLC (E). (C) Western blot of adult fluke secretions probed with an anti-FhHDM-1 antibody. P, pre-immune sera; T, test bleed. (D) N-terminal sequencing and LC-MS/MS analysis of the native ∼ 6 kDa protein generated peptide sequence information that allowed cloning of the cDNA, termed FhHDM-1. The primary amino acid sequence of FhHDM-1 derived from conceptual translation of the cDNA is shown. The predicted N-terminal signal peptide is shown in italics and the actual N-terminal of the native protein is shown by an arrow. The SEESREKLRE sequence generated by N-terminal sequencing is boxed in grey and a peptide (m/z 642.93; ITEVITILLNR) matched by LC-MS/MS following tryptic digest of the native protein is underlined. Secondary structure predictions using using PSIPRED [26], shown below the primary sequence, suggest the molecule is predominantly α-helical.
