The MF6p/FhHDM-1 major antigen secreted by the trematode parasite Fasciola hepatica is a heme-binding protein [Protein Structure and Folding]
November 26th, 2013 by Martinez-Sernandez, V., Mezo, M., Gonzalez-Warleta, M., Perteguer, M. J., Muino, L., Guitian, E., Garate, T., Ubeira, F. M.
Blood-feeding parasites have developed bio-chemical mechanisms to control heme intake and detoxification. Here we show that a major antigen secreted by Fasciola hepatica, previously reported as MF6p, of unknown function (gb|CCA61804.1), and as FhHDM-1, considered to be a helminth defence molecule belonging to the family of cathelicidin-like proteins (gb|ADZ24001.1), is in fact a heme-binding protein. The heme-binding nature of the MF6p/FhHDM-1 protein was revealed in two independent experiments: i) immunopurifica-tion of the secreted protein:heme complexes with mAb MF6, and subsequent analysis by C8 reversed-phase HPLC and MS/MS spectrometry, and ii) analysis of the binding ability of the synthetic protein to hemin in vitro. By immunohistochemistry analysis we have observed that MF6p/FhHDM-1 is pro-duced by parenchymal cells and transported to other tissues (e.g., vitellaria and testis). Interestingly, MF6p/FhHDM-1 is absent in both the intestinal cells and in the lumen of caecum, but can be released through the tegumental surface to the external medium, where it binds to free heme molecules regurgitated by the parasite after hemoglobin digestion. Proteins that are close analogs of the Fasciola MF6p/FhHDM-1 are present in other trematodes, including Clonorchis, Opistorchis, Paragonimus, Schistosoma and Dicrocoelium. Using UV-visible spectroscopy and im-munoprecipitation techniques, we observed that synthetic MF6p/FhHDM-1 binds to hemin with 1:1 stoichiometry and an apparent Kd of 1.14x10-6 M-1. We also demonstrated that for-mation of synthetic MF6p/FhHDM-1:hemin complexes inhibited hemin degradation by hydrogen peroxide, and hemin peroxidase-like activity in vitro. Our results suggest that MF6p/FhHDM-1 may be involved in heme homeostasis in trematodes.