Aspartic peptidases of the pinewood nematode Bursaphelenchus xylophilus: Molecular characterization and in silico structural analysis

Aspartic peptidases, a family of proteinases with catalytic aspartate residues in the active site, have been reported in a large variety of organisms associated with different functions in animal parasitism. However, scarce studies have been performed on plant-parasitic nematode aspartic peptidases and their possible role in parasitism. The pinewood nematode, Bursaphelenchus xylophilus, is an important plant-parasitic nematode responsible for the pine wilt disease and recognized as a major forest pest. In the present study, the cDNA coding sequence of three aspartic peptidases secreted by B. xylophilus (BxASP101, BxASP102 and BxASP103) was cloned and sequenced to predict complete amino acid sequences. Phylogenetic analysis indicated that the three peptidases clearly differ from other nematode aspartic peptidases, forming a separate branch. In silico three-dimensional structure analysis predicted monomers with bilobal configuration and an extended active site cleft localized between the two lobes, with each N-and C-terminal lobe contributing with one active aspartic acid residue. The gene transcript levels for these three proteins were higher in B. xylophilus stimulated with Pinus pinaster extract, a high-susceptible tree, than when stimulated with P. pinea extract, a low-susceptible tree. These results revealed that BxASP101, BxASP102 and BxASP103 are aspartic peptidases, which might be related to B. xylophilus pathogenicity and can be further explored to develop new target-specific strategies for the management of this nematode.