Structural and functional investigation of a secreted chorismate mutase from the plant-parasitic nematode Heterodera schachtii in the context of related enzymes from diverse origins.
In this article, we present the cloning of Hscm1, a gene for chorismate mutase (CM) from the beet cyst nematode Heterodera schachtii. CM is a key branch-point enzyme of the shikimate pathway, and secondary metabolites that arise from this pathway control developmental programmes and defence responses of the plant. By manipulating the plant's endogenous shikimate pathway, the nematode can influence the plant physiology for its own benefit. Hscm1 is a member of the CM gene family and is expressed during the pre-parasitic and parasitic stages of the nematode's life cycle. In situ mRNA hybridization reveals an expression pattern specific to the subventral and dorsal pharyngeal glands. The predicted protein has a signal peptide for secretion in addition to two domains. The N-terminal domain of the mature protein, which is only found in cyst nematodes, contains six conserved cysteine residues, which may reflect the importance of disulphide bond formation for protein stabilization. The C-terminal domain holds a single catalytic site and has similarity to secreted CMs of pathogenic bacteria, classifying HsCM1 as an AroQ(gamma) enzyme. The presumed catalytic residues are discussed in detail, and genetic complementation experiments indicate that the C-terminal domain is essential for enzyme activity. Finally, we show how the modular design of the protein is mirrored in the genomic sequence by the intron/exon organization, suggesting exon shuffling as a mechanism for the evolutionary assembly of this protein.
Vanholme
Bartel
B
Molecular Biotechnology Department, Faculty of Bioscience Engineering, Ghent University, Coupure links 653, B-9000 Ghent, Belgium.
Kast
Peter
P
Haegeman
Annelies
A
Jacob
Joachim
J
Grunewald
Wim
W
Gheysen
Godelieve
G
D000595:Amino Acid Sequence; D000818:Animals; D015139:Blotting, Southern; D002826:Chorismate Mutase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005057:Eukaryotic Cells / Q000201:enzymology; D018507:Gene Expression Regulation, Developmental; D017238:Genes, Helminth; D005816:Genetic Complementation Test; D015801:Helminth Proteins / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D010271:Parasites / Q000201:enzymology; D010802:Phylogeny; D010944:Plants / Q000469:parasitology*; D011387:Prokaryotic Cells / Q000201:enzymology; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020539:Sequence Analysis, Protein; D014431:Tylenchoidea / Q000201:enzymology* / Q000235:genetics
D015801:Helminth Proteins; D002826:Chorismate Mutase