Crystal structure of an extracellular superoxide dismutase from Onchocerca volvulus and implications for parasite-specific drug development

Abstract

Superoxide dismutases (SODs) are metalloproteins that are responsible for the dismutation of superoxide anion radicals. SODs are consequently protective against oxidative damage to cellular components. Among other protective mechanisms, the filarial parasite Onchocerca volvulus has a well developed defense system to scavenge toxic free radicals using SODs during migration and sojourning of the microfilariae and adult worms in the human body. O. volvulus is responsible for the neglected disease onchocerciasis or `river blindness'. In the present study, an extracellular Cu/Zn-SOD from O. volvulus (OvEC-SOD) was cloned, purified and crystallized to obtain structural insight into an attractive drug target with the potential to combat onchocerciasis. The recombinant OvEC-SOD forms a dimer and the protein structure was solved and refined to 1.55 Å resolution by X-ray crystallography. Interestingly, a sulfate ion supports the coordination of the conserved copper ion. The overall protein shape was verified by small-angle X-ray scattering. The enzyme shows a different surface charge distribution and different termini when compared with the homologous human SOD. A distinct hydrophobic cleft to which both protomers of the dimer contribute was utilized for a docking approach with compounds that have previously been identified as SOD inhibitors to highlight the potential for individual structure-based drug development.

Keywords: Cu/Zn superoxide dismutases; Onchocerca volvulus; X-ray crystallography; docking; drug targets; metal ion coordination; parasites.

Figure 1

High-resolution crystal structure of OvEC-SOD. (a) Topology diagram of the protein and its secondary structure. (b) Superposition of O. volvulus SOD (green) with T. solium SOD (pink; PDB entry 3mnd; C^α r.m.s.d. of 0.5 Å; Hernández-Santoyo et al., 2011 ▸) and H. sapiens SOD (purple); PDB entry 1hl4; C^α r.m.s.d. of 0.5 Å; Strange et al., 2003 ▸). (c) Cartoon and surface representation of the crystallographic OvEC-SOD dimer. The approximate position of the active site as highlighted in (d) is framed. (d) Close-up of the active site and the two conserved metal ions. The coordination distances are shown in Å.

Figure 2

Solution structure of OvEC-SOD. (a) Averaged normalized scattering intensities (blue) plotted against the momentum transfer s as obtained by SAXS. The calculated fit curve (green) is a comparison with the ab initio model shown as gray spheres in (c). (b) Scattering intensities and fit curve as shown in (a) superposed with additional fit curves of the OvEC-SOD dimer crystal structure with the SAXS data using either CRYSOL (magenta) or SREFLEX (yellow) for structural comparison. (c) Superposition of the OvEC-SOD dimer crystal structure in green and the ab initio model of OvEC-SOD based on the SAXS data.

Figure 3

Docking of LeadQuest compounds to OvEC-SOD (gray): (a) ID 1545-7806 (molecular mass 426.5 g mol⁻¹; C₂₃H₂₆N₂O₄S), (b) ID 1460-00055 (molecular mass 562.6 g mol⁻¹; C₃₀H₂₈F₆N₂O₂), (c) ID 1502-3317 (molecular mass 438.6 g mol⁻¹; C₇₇H₈₇F₆N₁₁O₁₀S).