Characterization of two cDNAs encoding cysteine proteinases from the soybean cyst nematode Heterodera glycines.
Two cDNAs encoding cysteine proteinases were isolated from a cDNA library constructed from feeding females of Heterodera glycines. The library was screened with a cysteine proteinase gene fragment originally amplified from cDNA of H. glycines. Database searches predict that 1 cDNA (hgcp-I) encodes a cathepsin L-like proteinase, while the second (hgcp-II) encodes a cathepsin S-like enzyme. Both predicted proteins contain a short secretion signal sequence, a long propeptide and a mature protein of 219 amino acids. Southern blot analysis suggests that the cathepsin S-like enzyme, HGCP-II, is encoded by a single-copy gene in contrast to the cathepsin L-like proteinase, HGCP-I which may have 2 homologues. The regions encoding the mature proteinases were cloned into an expression vector and recombinant protein produced in E. coli. HGCP-I was shown, after refolding, to cleave the synthetic peptide Z-Phe-Arg-AMC, and this activity could be inhibited by the engineered rice cystatin Oc-I delta D86. HGCP-II showed no activity against the synthetic substrates tested. The knowledge gained from these studies will improve our understanding of plant nematode proteinases and aid the development of a rational proteinase inhibitor-based approach to plant nematode resistance.
Urwin
P E
PE
Centre for Plant Biochemistry and Biotechnology, University of Leeds, UK.
Lilley
C J
CJ
McPherson
M J
MJ
Atkinson
H J
HJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000235:genetics* / Q000378:metabolism; D018509:DNA, Helminth; D005260:Female; D015870:Gene Expression; D015801:Helminth Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009348:Nematoda / Q000201:enzymology* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D013025:Glycine max / Q000469:parasitology*
D018509:DNA, Helminth; D015801:Helminth Proteins; D011993:Recombinant Fusion Proteins; D003546:Cysteine Endopeptidases