Purification and characterization of a 16-kDa cysteine proteinase of Gymnophalloides seoi (Gymnophallidae) metacercariae.
Gymnophalloides seoi, a new human intestinal trematode transmitted by oysters, is highly prevalent in southwestern coastal areas of Korea. The aim of this preliminary study was to acquire an understanding of the pathogenesis of G. seoi infection, and, to this end, we followed 2 consecutive steps, Sephacryl S-300 HR and CM-Trisacryl M chromatography, to purifiy a 16-kDa proteinase from crude extract of G. seoi metacercariae. Enzyme activities were completely inhibited by L-trans-epoxysuccinylleucylamide (4-guanidino) butane (E-64) and iodoacetic acid, cysteine proteinase inhibitors, strongly suggesting that the enzyme is a cysteine proteinase. Activity of the proteinase was maximal at pH 5.0 in 0.1 M of buffer and potentiated in the presence of 5 mM dithiothreitol. The proteinase showed differential abilities to hydrolyze macromolecules and immunoglobulins; it completely degraded extracellular matrix proteins such as collagen and fibronectin during overnight incubation but digested hemoglobin very slowly. The proteinase also cleaved human immunoglobulins IgG2a and sIgA; heavy chains were more susceptible than light chains to its digestive activity. The proteinase showed no antigenicity on both enzyme-linked immunosorbent assay and immunoblots, however. Our results strongly suggest that the cysteine proteinase of G. seoi metacercariae is potentially significant in nutrient uptake and evasion by the worm of the host immune response.
Choi
M H
MH
Department of Parasitology and Institute of Endemic Diseases, Seoul National University College of Medicine, Korea.
Chai
J Y
JY
Lee
S H
SH
D000818:Animals; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003094:Collagen / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005353:Fibronectins / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D007136:Immunoglobulins / Q000378:metabolism; D010125:Ostreidae; D011480:Protease Inhibitors / Q000494:pharmacology; D014200:Trematoda / Q000201:enzymology*
D005353:Fibronectins; D006454:Hemoglobins; D007136:Immunoglobulins; D011480:Protease Inhibitors; D003094:Collagen; D003546:Cysteine Endopeptidases