title: Structural and functional characterization of a novel, host penetration-related pectate lyase from the potato cyst nematode Globodera rostochiensis.
abstract: SUMMARY The cell wall, a strong extraprotoplasmic layer surrounding plant cells that mainly consists of a variety of polysaccharides, constitutes a major barrier for potential parasites. Plant-parasitic nematodes are well equipped to overcome this barrier as they produce and secrete cell-wall-degrading enzymes. Expression profiling of various life stages of the potato cyst nematode Globodera rostochiensis revealed a novel pectate lyase gene (Gr-pel2, 759 bp). The Gr-PEL2 protein showed highest similarity to pectate lyases from the facultative plant-parasitic nematodes Bursaphelenchus mucronatus and B. xylophilus and the soil-inhabiting saprophytic Streptomyces and Frankia species (i.e. 40-42% identity and 58-60% similarity), whereas only a remote relatedness to the previously identified Gr-PEL1 was observed (i.e. 28% identity and 43% similarity). Transient expression of Gr-pel2 in leaves of Nicotiana benthamiana resulted in severe malformations of the infiltrated tissues, not relating to maceration and soft rot symptoms. Ca(2+) is known to be essential for pectate lyase activity, and the most likely calcium-binding site was identified in the Gr-PEL2 protein by combining homology modelling of the three-dimensional structure, site-directed mutagenesis and transient expression in leaves. A highly charged cleft in Gr-PEL2, which is likely to be involved in substrate binding and which is also significantly more hydrophobic in Gr-PEL1, was shown to be essential for protein activity. Our results underline the broad spectrum of pectate lyases and cell-wall-degrading enzymes necessary for successful parasitism by cyst nematodes.
authors.affiliation: Laboratory of Nematology, Wageningen University, Binnenhaven 5, 6709 PD Wageningen, the Netherlands.
authors.forename: Urszula
authors.initials: U
authors.lastname: Kudla
authors.forename: Adina-Luminita
authors.initials: AL
authors.lastname: Milac
authors.forename: Ling
authors.initials: L
authors.lastname: Qin
authors.forename: Hein
authors.initials: H
authors.lastname: Overmars
authors.forename: Erwin
authors.initials: E
authors.lastname: Roze
authors.forename: Martijn
authors.initials: M
authors.lastname: Holterman
authors.forename: Andrei-Jose
authors.initials: AJ
authors.lastname: Petrescu
authors.forename: Aska
authors.initials: A
authors.lastname: Goverse
authors.forename: Jaap
authors.initials: J
authors.lastname: Bakker
authors.forename: Johannes
authors.initials: J
authors.lastname: Helder
authors.forename: Geert
authors.initials: G
authors.lastname: Smant