A new multi-domain member of the cystatin superfamily expressed by Fasciola hepatica.
Cystatins are cysteine protease inhibitors that are widespread in the plant and animal kingdoms. Cystatins are expressed by helminth parasites that may employ these proteins to regulate parasite cysteine protease activity and to modulate host immune responses. Here, we describe the cloning of a cDNA encoding a high molecular weight protein of Fasciola hepatica that contains two domains with significant identity to the cardinal cystatin signatures and four domains with degenerated cystatin signatures. This is the first report of a multi-domain cystatin in an invertebrate species. While cystatins are divided into three evolutionary related families, our phylogenetic analysis shows that all cystatin domains within this protein, like several other helminth cystatins, belong to the cystatin family 2. The DNA region encoding the domain 4 that is the best conserved at the level of its cystatin signatures was expressed in Drosophila cells and a recombinant protein was produced and purified. This protein was a potent inhibitor of the papain and of the major cysteine protease of F. hepatica, the cathepsin L1.
Khaznadji
Eric
E
INRA, UR86 Bio-Agresseurs, Santé et Environnement 37 380 Nouzilly, France.
Collins
Peter
P
Dalton
John P
JP
Bigot
Yves
Y
Moiré
Nathalie
N
D000595:Amino Acid Sequence; D000818:Animals; D015139:Blotting, Southern / Q000379:methods; D002403:Cathepsins / Q000037:antagonists & inhibitors; D003001:Cloning, Molecular / Q000379:methods; D015891:Cystatins / Q000235:genetics* / Q000302:isolation & purification; D015853:Cysteine Proteinase Inhibitors / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D005210:Fasciola hepatica / Q000235:genetics*; D015870:Gene Expression / Q000235:genetics; D017238:Genes, Helminth / Q000235:genetics; D015801:Helminth Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D010206:Papain / Q000037:antagonists & inhibitors; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification; D016415:Sequence Alignment / Q000379:methods
D015891:Cystatins; D015853:Cysteine Proteinase Inhibitors; D018076:DNA, Complementary; D015801:Helminth Proteins; D011994:Recombinant Proteins; D002403:Cathepsins; C509464:cathepsin L1, Fasciola hepatica; D010206:Papain