FhCaBP4: a Fasciola hepatica calcium-binding protein with EF-hand and dynein light chain domains

Abstract

In trematodes, there is a family of proteins which combine EF-hand-containing domains with dynein light chain (DLC)-like domains. A member of this family from the liver fluke, Fasciola hepatica-FhCaBP4-has been identified and characterised biochemically. FhCaBP4 has an N-terminal domain containing two imperfect EF-hand sequences and a C-terminal dynein light chain-like domain. Molecular modelling predicted that the two domains are joined by a flexible linker. Native gel electrophoresis demonstrated that FhCaBP4 binds to calcium, manganese, barium and strontium ions, but not to magnesium or zinc ions. The hydrophobic, fluorescent probe 8-anilinonaphthalene-1-sulphonate bound more tightly to FhCaBP4 in the presence of calcium ions. This suggests that the protein undergoes a conformational change on ion binding which increases the number of non-polar residues on the surface. FhCaBP4 was protected from limited proteolysis by the calmodulin antagonist W7, but not by trifluoperazine or praziquantel. Protein-protein cross-linking experiments showed that FhCaBP4 underwent calcium ion-dependent dimerisation. Since DLCs are commonly dimeric, it is likely that FhCaBP4 dimerises through this domain. The molecular model reveals that the calcium ion-binding site is located close to a key sequence in the DLC-like domain, suggesting a plausible mechanism for calcium-dependent dimerisation.

FhCaBP1 (FH22): A Fasciola hepatica calcium-binding protein with EF-hand and dynein light chain domains

“…2a). This is similar to the predicted structures of the other FhCaBP and SmTAL family members (Banford, et al, 2013, Orr, et al, 2012. In common with the other family members, the region linking the two domains is expected to be flexible, allowing the two domains to alter their relative orientation.…”

“…We present the biochemical characterisation of FH22 (including predicted three-dimensional structure, ion binding, oligomerisation properties and interactions with calmodulin antagonists), building on previously reported work on this protein and complementing our own work on other family members (Ruiz de Eguino, et al, 1999) (Banford, et al, 2013, Nguyen, et al, 2016, Orr, et al, 2012. We report data on the protein's ion and drug binding properties, predicted structure and dimerization.…”

“…Thus, like other CaBP family members, FhCaBP1 can interact with this ion in addition to calcium. FhCaBP3 and FhCaBP4 both bind other ions in addition to calcium and manganese; however, like FhCaBP1, FhCaBP2 only binds these two ions (Banford, et al, 2013, Orr, et al, 2012). FhCaBP1's ability to bind calcium ions was confirmed by intrinsic fluorescence measurements and differential scanning fluorimetry.…”

“…However in this case, calcium ion binding promotes the monomeric over the dimeric state (which is favoured in the absence of these ions). Interestingly, in FhCaBP4 the situation is reversed: calcium ion binding promotes dimerization (Orr, et al, 2012). All three of these proteins interact with calmodulin antagonists and related drugs; however, the range of drugs which binds varies between the proteins, demonstrating that they can be distinguished pharmacologically (Banford, et al, 2013, Orr, et al, 2012.…”

Molecular interactions of proteins associated with surface membrane biogenesis and renewal in the tegument of Schistosoma mansoni

“…Among these families, the TALs are of particular interest for schistosomiasis and other trematodiases. TAL homologues have been found in the tegument of Fasciola gigantica (Subpipattana et al, 2012;Vichasri-Grams et al, 2006), Fasciola hepatica (Banford et al;Orr et al, 2012), Opisthorchis viverrini (Mulvenna et al, 2010b) and Clonorchis sinensis (Chen et al, 2011;Huang et al, 2007;Kim et al, 2012;Zhou et al, 2007). The presence of TAL family members in the tegument of a variety of helminths suggests that these molecules are important for the host immune response to helminths.…”