title: Characterization of intestinally active proteinases of cyst-nematodes.
abstract: Cryostat sections of juvenile and adult female stages of the soybean cyst-nematode, Heterodera glycines, were incubated with 4 different naphthylamide-linked peptide substrates to localize and characterize proteinase activity within the animal. Detected activity was restricted to the intestine and 2 distinct classes of proteinase were identified on the basis of substrate specificity and sensitivity to plant proteinase inhibitors. A cathepsin L-like cysteine proteinase activity capable of hydrolysing the synthetic substrates Z-Ala-Arg-Arg-MNA and Z-Phe-Arg-MNA but not Z-Arg-Arg-MNA or L-Arg-NA was inhibited by an engineered variant of a cysteine proteinase inhibitor from rice (Oc-I delta D86). The cleavage of Z-Phe-Arg-MNA was sensitive to inhibition by a combination of Oc-I delta D86 and cowpea trypsin inhibitor (CpTI). Degenerate oligonucleotide primers were used to amplify fragments of cysteine proteinase genes from 2 cyst-nematodes, H. glycines and Globodera pallida. Comparison of the H. glycines fragment with known genes established highest homology to cathepsin L-like genes. In contrast, the amplified G. pallida fragment displayed greatest homology to cathepsin B-like genes from Caenorhabditis elegans.
authors.affiliation: Centre for Plant Biochemistry and Biotechnology, University of Leeds, UK.
authors.forename: C J
authors.initials: CJ
authors.lastname: Lilley
authors.forename: P E
authors.initials: PE
authors.lastname: Urwin
authors.forename: M J
authors.initials: MJ
authors.lastname: McPherson
authors.forename: H J
authors.initials: HJ
authors.lastname: Atkinson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015891:Cystatins / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D018509:DNA, Helminth; D005260:Female; D007422:Intestines / Q000201:enzymology; D008969:Molecular Sequence Data; D009348:Nematoda / Q000201:enzymology*; D010944:Plants / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism*
chemical_list: D015891:Cystatins; D015853:Cysteine Proteinase Inhibitors; D018509:DNA, Helminth; D012697:Serine Endopeptidases; D003546:Cysteine Endopeptidases