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European Biophysics Journal

, Volume 32, Issue 5, pp 465–476 | Cite as

The polyprotein and FAR lipid binding proteins of nematodes: shape and monomer/dimer states in ligand-free and bound forms

  • Alexandra S. Solovyova
  • Nicola Meenan
  • Lindsay McDermott
  • Antonio Garofalo
  • Jannette E. Bradley
  • Malcolm W. Kennedy
  • Olwyn Byron
Article

Abstract

Nematodes produce two classes of small, helix-rich fatty acid- and retinol-binding proteins whose structures and in vivo functions remain to be elucidated. These are the polyprotein allergens (NPA) and the FAR proteins. The solution properties of recombinant forms of these proteins from parasitic [Ascaris suum (As) and Onchocerca volvulus (Ov)] and free-living [Caenorhabditis elegans (Ce)] nematodes have been examined. Analytical ultracentrifugation (AUC) showed that, contrary to previous findings, the rAs-NPA-1A polyprotein unit (~15 kDa) is a monomer, and this stoichiometry is unaltered by ligand (oleic acid) binding. The rOv-FAR-1 and rCe-FAR-5 proteins differ in that the former forms a tight dimer and the latter a monomer, and these oligomeric states are also unaffected by ligand binding or protein concentration. Sedimentation equilibrium experiments showed that the partial specific volume v̄ of the unliganded proteins agree well with the value calculated from amino acid composition extrapolated to experimental temperature, and was unaffected upon ligand binding. Data from small-angle X-ray scattering (SAXS) indicated that both of the monomeric proteins rAs-NPA-1A and rCe-FAR-5 are globular, although slightly elongated and flattened. These data are in good agreement with shapes predicted from sedimentation velocity experiments and hydrodynamic bead modelling. On the basis of functional and secondary structural homology with the ligand-binding domain of the retinoic acid receptor RXRα, de novo atomic resolution structures for rAs-NPA-1A and rCe-FAR-5 have been constructed which are consistent with the SAXS and hydrodynamic data.

Keywords

Analytical ultracentrifugation Hydrodynamic bead modelling Nematode fatty acid binding proteins Protein-ligand interaction Small-angle X-ray scattering 

Notes

Acknowledgements

A.S.S. is supported by a grant from the Wellcome Trust to O.B., M.W.K. and others (05606/Z/99/Z), and we are also indebted to the Wellcome Trust for their support of this project through other grants to M.W.K. The work was also supported by an ESRF beam time award (LS-1872) and by the Universities of Salford and Nottingham. Special thanks to the team of Dr. Wim Bras (ESRF, Grenoble, particularly Dr. Marc Malfois) for assistance in the SAXS experiments and Marcelo Nöllmann for help in SAXS data treatment.

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Copyright information

© EBSA 2003

Authors and Affiliations

  • Alexandra S. Solovyova
    • 1
    • 2
  • Nicola Meenan
    • 3
  • Lindsay McDermott
    • 3
    • 4
  • Antonio Garofalo
    • 5
  • Jannette E. Bradley
    • 5
  • Malcolm W. Kennedy
    • 4
  • Olwyn Byron
    • 1
  1. 1.Division of Infection and Immunity, Institute of Biomedical and Life Sciences, Joseph Black BuildingUniversity of GlasgowGlasgow UK
  2. 2.Institute for Problems of Cryobiology and CryomedicineNational Academy of Science of the UkraineKharkovUkraine
  3. 3.Department of Chemistry, Joseph Black BuildingUniversity of GlasgowGlasgow UK
  4. 4.Division of Environmental and Evolutionary Biology, Institute of Biomedical and Life Sciences, Graham Kerr BuildingUniversity of GlasgowGlasgow UK
  5. 5.School of Life and Environmental ScienceUniversity of NottinghamNottinghamUK

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