Molecular characterization and functional analysis of a pathogenesis-related β-1,3-glucanase gene in spruce (Picea asperata).

β-1,3-glucanase, a member of the plant PR-2 family of pathogenesis-related (PR) proteins and Glycoside Hydrolase Family 17 (GH17), plays an important role in plant defense mechanisms against pathogens. In this study, a β-1,3-glucanase gene from Picea asperata was isolated and designated as PaGlu; the gene consisted of a 1035 bp open reading frame (ORF) encoding 344 amino acid residues. Multiple alignment and phylogenetic analysis showed that PaGlu has the (91.86%) highest identity and the closest relationship with the β-1,3-glucanase of Picea sitchensis. qRT-PCR analysis revealed that the PaGlu gene was mainly expressed in the needles of Pheidole asperata. Furthermore, the expression of the PaGlu gene was significantly upregulated by L. piceae, suggesting that it is involved in the defense response of P. asperata against L. piceae infection. The optimal induction condition for recombinant PaGlu in Escherichia coli was incubation with 0.6 mM IPTG (isopropyl β-D-thiogalactoside) for 6 h at 25 °C, and the molecular weight of recombinant PaGlu was determined to be approximately 53 kDa. The purified recombinant PaGlu displayed high enzyme activity (20.7 U/mg) and the optimal conditions were pH 6.0 and 50 °C. In addition, the purified recombinant PaGlu exhibited a different destructive effect on the mycelium of Lophodermium piceae, Colletotrichum gloeosporioides, Fusarium proliferatum and Fusarium sporotrichioides. Importantly, it was able to destroy the hyphal wall of L. piceae. These results provide a molecular basis for further understanding of the mechanisms of the PaGlu gene in P. asperata against L. piceae infection. [ABSTRACT FROM AUTHOR]