Expression and functional analysis of polygalacturonase gene Pcipg5 from the plant pathogen Phytophthora capsici

Abstract

As one of the cell wall-degrading enzymes, polygalacturonase involves in pathogenicity and virulence in a number of host pathogen interactions. In the polygalacturonase multigene family of Phytophthora capsici , Pcipg5 was screened from the genomic library. The Pcipg5 gene contains three putative active sites (179Asp, 200Asp, 201Asp) and one potential N-glycosylation site (252Asn). PCIPG5, four active-site mutagenesis proteins and N-glycosylation mutation were expressed in Pichia pastoris and purified by Ni-NTA Purification System. Pcipg5 functions were determined by Western blot, RT-PCR and Northern blot. Transmission electron microscopy discovery individually in treated pepper leaves was used to determine the virulence of PCIPG5. The results show that Pcipg5 can encode a pathogenesis-related protein during P. capsici infection of pepper leaves and degrade cell walls to produce necrotic lesions in treated pepper leaves. Meanwhile, the N-glycosylation mutagenesis protein decreased the activity compared with PCIPG5. It indicates that the existence N-glycosylation site in PCIPG5 plays a partial role in the activity of this enzyme. Key words : Phytophthora capsici, Pcipg5, host-pathogen interactions, N-glycosylation.