Characterisation of a novel Kunitz-type molecule from the trematode Fasciola hepatica.
A low molecular mass monomeric protein termed Fh-KTM (Fasciola hepatica Kunitz-type molecule) was isolated from the trematode Fasciola hepatica. Fh-KTM is a single polypeptide of 58 amino acids and a Mr of 6751. The complete amino acid sequence of Fh-KTM was determined and revealed significant similarity to the Kunitz-type (BPTI) family of proteinase inhibitors. Several polymorphisms were observed suggesting that more than one Fh-KTM molecule may be expressed by this parasite. Modified proline residues were shown to occur at all four positions in this protein as 3-hydroxy derivatives. This is the first report of 3-hydroxyproline residues in a Kunitz-type molecule. Indirect immunofluorescence and immunogold labelling revealed that Fh-KTM is an abundant molecule within the parasite localised to the gut, the parenchymal tissue and the tegument of adult F. hepatica. Serine protease inhibition assays revealed that Fh-KTM exhibited little or no inhibition against chymotrypsin, kallikrein, urokinase or key serine proteases of the blood coagulation pathways. However, Fh-KTM was able to inhibit trypsin even though the P1 reactive amino acid of Fh-KTM was a leucine residue.
Bozas
S E
SE
Immunoparasitology Department, Victorian Institute of Animal Science, Victoria, Australia. bozase@woody.agvic.gov.au
Panaccio
M
M
Creaney
J
J
Dosen
M
M
Parsons
J C
JC
Vlasuk
G V
GV
Walker
I D
ID
Spithill
T W
TW
D000595:Amino Acid Sequence; D000818:Animals; D007611:Aprotinin / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D005210:Fasciola hepatica / Q000737:chemistry / Q000235:genetics*; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D015842:Serine Proteinase Inhibitors / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D014361:Trypsin Inhibitors / Q000737:chemistry / Q000235:genetics
D015801:Helminth Proteins; D015842:Serine Proteinase Inhibitors; D014361:Trypsin Inhibitors; D007611:Aprotinin