Characterization of excretory–secretory products from protoscoleces of Echinococcus granulosus and evaluation of their potential for immunodiagnosis of human cystic echinococcosis (original) (raw)
Related papers
International Journal For Parasitology, 1992
KuMAR S. and PRITCHARD D. I. 1992. The partial characterization of proteases present in the excretory/secretory products and exsheathing fluid of the infective (L3) larva of Necator americanus. International Journal for Parasitology 22: 563-572. Following the observation that live third-stage larvae (L3) could digest gelatin in vitro, gelatinolytic protease activity has been demonstrated at pH 8.5, in both exsheathing fluid (EF) and excretory/secretory (ES) products of infective L3 of Necator americanus. EF resolved as a single band of proteolytic activity, with a mol. wt of 116 kDa, while L3 ES products exhibited multiple bands ofproteolysis, at 219,200, 195, 166,137,92,72 and 62 kDa; weak bands were detectable at 92 and 72 kDa. The EF protease was characterized as cysteine, whereas ES apparently possessed one serine (195 kDa) and seven (219, 200, 166, 137, 92, 72 and 62 kDa) cysteine protease bands and a combination of metallo-and cysteine proteases of approximately the same mol. wts (62, 137 and 219 kDa). Though EF was not able to cleave immunoglobulins, ES was shown to cleave IgG, IgA and IgM, but not IgD or IgE. The activity appeared to be directed toward the Fc portion of the molecule, and was inhibited by PMSF, which is indicative of serine protease activity. The significance of the presence of such apparently diverse proteases in larval products is discussed.
1980
Three alkaline proteases from the larval gut of Spodoptera litura have been purified by gel filtration on Sephadex G-75 and ion exchange chromatography on DEAE-Sephadex A-50. Optimal pH for the activities of enzymes using casein as substrate were 11.0, 10.5 and 9.0 respectively. Proteases I-III have estimated molecular weight values of 17,000, 21,000 and 53,000 when determined by gel filtration and 18,000, 23,000 and 50,000 when determined by SDS polyacrylamide gel electrophoresis. All the enzymes were inhibited by phenylmethylsulphonyl fluoride suggesting the presence of serine in the active site. Sulphydryl reagents and metal chelating agent had no effect on the activity of any of the proteases. N<t-p-tosyl-L-lysine chloromethyl ketone HCI inhibited the activity of all three enzymes indicating the participation of histidine in the active site whereas L-l-tosylamide-2-phenylethyl chloromethyl ketone showed no inhibitory effect of any of the enzymes. Mg 2+, Ca 2+, Mn 2+ and Co 2÷ had no effect on the activity of the enzymes, however, Cu 2 +, Zn 2 ÷ and Hg 2÷ caused inhibition. The K values as determined with casein substrate were 5.7 x 10-6M, 2.9 x 10-6M, and 2.1 x 10-6M for proteinase I-III, respectively. A comparison is made between S. litura proteases, other insect proteases and the better known mammalian proteases.
Insect Biochemistry, 1980
Three alkaline proteases from the larval gut of Spodoptera litura have been purified by gel filtration on Sephadex G-75 and ion exchange chromatography on DEAE-Sephadex A-50. Optimal pH for the activities of enzymes using casein as substrate were 11.0, 10.5 and 9.0 respectively. Proteases I-III have estimated molecular weight values of 17,000, 21,000 and 53,000 when determined by gel filtration and 18,000, 23,000 and 50,000 when determined by SDS polyacrylamide gel electrophoresis. All the enzymes were inhibited by phenylmethylsulphonyl fluoride suggesting the presence of serine in the active site. Sulphydryl reagents and metal chelating agent had no effect on the activity of any of the proteases. N<t-p-tosyl-L-lysine chloromethyl ketone HCI inhibited the activity of all three enzymes indicating the participation of histidine in the active site whereas L-l-tosylamide-2-phenylethyl chloromethyl ketone showed no inhibitory effect of any of the enzymes. Mg 2+, Ca 2+, Mn 2+ and Co 2÷ had no effect on the activity of the enzymes, however, Cu 2 +, Zn 2 ÷ and Hg 2÷ caused inhibition. The K values as determined with casein substrate were 5.7 x 10-6M, 2.9 x 10-6M, and 2.1 x 10-6M for proteinase I-III, respectively. A comparison is made between S. litura proteases, other insect proteases and the better known mammalian proteases.
Antibody response to a protease secreted byTrichinella spiralis muscle larvae
Parasitology Research, 1995
In the present study we analyzed the humoral response of Trichinella spiralis-infected mice to a 35-kDa protease (purified from the excretory-secretory products of T. spiralis muscle larvae) by a Western-blot procedure and an enzyme-linked immunosorbent assay (ELISA) technique using a panel of postinfection mouse anti-Trichinella sera. The results demonstrated that this response was time-dependent and that infected mice could be distinguished from controls. In addition, inhibition assays demonstrated that these antisera were capable of abolishing the proteinase activity of the 35-kDa protease in vitro. The occurrence of proteases seems to be a very common feature in parasite crude extracts and excretory-secretory products (McKerrow 1989). It is also known that these enzymes are implicated in important host-parasite interactions, and for this reason, recent reports have proposed the use of parasite proteases both as alternative targets for an induced immune response and as a rich source of antigenic material for diagnostic testing (
Insect Biochemistry, 1987
Al~ract--Proteinases contained in the posterior midgut of the stable fly, Stomoxys calcitrans (L.), have been tentatively identified by posterior midgut hydrolysis of synthetic substrates. Trypsin was identified by maximal hydrolysis of benzoyl-DL-arglnlne-p-nitroanilide (BAPNA) and tosyl-L-arginine methyl ester (TAME) at pH 8.0 and chymotrypsin, by maximal hydrolysis of benzoyl-L-tyrosine ethyl ester (BTEE) at pH 8.0. Carboxypeptidase A and B were identified by their maximal hydrolysis of hippuryl-DL-phenyllactic acid and hippuryl-L-arglnlne at pH 8.0 and 7.5 respectively and aminopeptidase was identified by maximal hydrolysis of leucine-p-nitroanilide at pH 7.5. Molecular weights, determined using gel exclusion chromatography, for proteinases were 19,500 for trypsin, 27,500 and 64,000 for chymotrypsin. Hydrolytic activity for the protein substrate haemoglobin occurred as two peaks with molecular weights of 19,500 and 27,500. Molecular weights for the peptidases were; carboxypeptidase A, 26,000, carboxypeptidase B, 28,500 and greater than 1,500,000 for aminopeptidase. These results indicate that the posterior midgut of the stable fly contains both proteinases and peptidases necessary for proteolytic breakdown of the ingested blood meal.
Alkaline protease in the larvae of the army worm, Spodoptera litura
Insect Biochemistry, 1976
The alkaline protease activity in the gut of Spodoptera litura was found to increase with the development of larvae and decreased with the onset of pupation. Fasting of the 5th instar larvae caused a slight increase in protease activity at 4 hr, which declined consistently on further starvation. The optimum pH for the gut protease was ll.0, with a shoulder between pH 8.0 and 9.0. The protease was inactivated upon dialysis of the crude enzyme solution at room temperature but not at 4°C. Incubation of the crude enzyme solution at pHI 1.0 and 37°C for 22 hr resulted in a three-fold rise in specific activity of the alkaline protease which declined on further incubation. The three-fold purified preparation, obtained by incubation of the crude enzyme solution, was passed through Sephadex G-75 to give a seven-fold purification and 70% yield, The preparation was not completely homogeneous and showed three clearly separable protein bands by polyacrylamide gel electrophoresis. The partially purified protease exhibited no shoulder between pH 8 to 9, like the crude preparation.
Zeitschrift für Naturforschung. C, Journal of biosciences, 2015
Proteins and proteolytic activities in the contents of the salivary gland (SGc), digestive tract (DTc) and excretory-secretory products (ESP) from larvae of the camel nasal botfly Cephalopina titillator were separated electrophoretically, and characterized. The protein profiles of the different samples were qualitatively quite similar in the larval stages L2 and L3. Zymogram analysis of proteases in the samples indicated that the digestive tract contained a greater variety of proteases than the salivary gland or the excretory-secretory products. They are mainly serine proteases. Proteases of ESP and DTc (especially of 3rd instar) contain trypsin- and chymotrypsin-like serine proteases, while the serine proteases of SGc are not of the trypsin- or chemotrypsin-type.
Cysteine proteases of parasitic organisms
Molecular and Biochemical Parasitology, 2002
Cysteine proteases play numerous indispensable roles in the biology of parasitic organisms. Aside from previously known general catabolic functions and protein processing, cysteine proteases may be key to parasite immunoevasion, excystment/encystment, exsheathing and cell and tissue invasion. Parasite cysteine proteases are unusually immunogenic and have been exploited as serodiagnostic markers and vaccine targets. Although host homologues exist, parasite cysteine proteases have distinct structural and biochemical properties including, pH optima and stability, the alteration in peptide loops or domain extensions, diverse substrate specificity and cellular location. The disparate nature of parasite cysteine protease compared to the host orthologous proteins has opened opportunities for chemotherapy. This review will highlight recent research on the 'papain-like' class of cysteine proteases, the most abundant family, and the newly discovered class of asparaginyl-endopeptidases. Cysteine protease classification will be re-examined in light of the diversity uncovered within parasitic organisms.
Parasites & vectors, 2017
Ticks, triatomines, mosquitoes and sand flies comprise a large number of haematophagous arthropods considered vectors of human infectious diseases. While consuming blood to obtain the nutrients necessary to carry on life functions, these insects can transmit pathogenic microorganisms to the vertebrate host. Among the molecules related to the blood-feeding habit, proteases play an essential role. In this review, we provide a panorama of proteases from arthropod vectors involved in haematophagy, in digestion, in egg development and in immunity. As these molecules act in central biological processes, proteases from haematophagous vectors of infectious diseases may influence vector competence to transmit pathogens to their prey, and thus could be valuable targets for vectorial control.