Anisakis simplex:Mutational Bursts in the Reactive Site Centers of Serine Protease Inhibitors from an Ascarid Nematode

Lu, Chuanyong; Nguyen, Thi Duyen; Morris, Sherri J.; Hill, Dolores E.; Sakanari, Judy

doi:10.1006/expr.1998.4284

Cited by 20 publications

(11 citation statements)

References 18 publications

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“…This group of low molecular weight serine protease inhibitors was first identified in A. suum and A. lumbricoides (16,18,51,54) where they are believed to protect the nematode from intestinal serine proteases. Members of the family have been identified in other parasitic nematodes, including the ascaridid A. simplex (20,52,56), the strongylid hookworms A. caninum and A. ceylanicum (17,23,25), and the whipworm T. suis (21,57). These SPIs are also believed to be involved in controlling exogenous host proteases in these nematodes.…”

Section: Discussionmentioning

confidence: 99%

Characterization of a Novel Filarial Serine Protease Inhibitor, Ov-SPI-1, from Onchocerca volvulus, with Potential Multifunctional Roles during Development of the Parasite

Ford

Guiliano

Oksov

et al. 2005

Journal of Biological Chemistry

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A novel filarial serine protease inhibitor (SPI) from the human parasitic nematode Onchocerca volvulus, Ov-SPI-1, was identified through the analysis of a molting third-stage larvae expressed sequence tag dataset. Subsequent analysis of the expressed sequence tag datasets of O. volvulus and other filariae identified four other members of this family. These proteins are related to the low molecular weight SPIs originally isolated from Ascaris suum where they are believed to protect the parasite from host intestinal proteases. The two Ov-spi transcripts are up-regulated in the molting larvae and adult stages of the development of the parasite. Recombinant Ov-SPI-1 is an active inhibitor of serine proteases, specifically elastase, chymotrypsin, and cathepsin G. Immunolocalization of the Ov-SPI proteins demonstrates that the endogenous proteins are localized to the basal layer of the cuticle of third-stage, molting third-stage, and fourth-stage larvae, the body channels and multivesicular bodies of third-stage larvae and the processed material found between the two cuticles during molting. In O. volvulus adult worms the Ov-SPI proteins are localized to the sperm and to eggshells surrounding the developing embryos. RNA interference targeting the Ov-spi genes resulted in the specific knockdown of the transcript levels of both Ov-spi-1 and Ov-spi-2, a loss of native proteins, and a significant reduction in both molting and viability of third-stage larvae. We suggest the Ov-SPI proteins play a vital role in nematode molting by controlling the activity of an endogenous serine protease(s). The localization data in adults also indicate that these inhibitors may be involved in other processes such as embryogenesis and spermatogenesis.The cuticle is an extracellular hydroskeleton that overlays the hypodermis of all nematodes. Most nematodes molt their cuticles four times during pre-adult development. Although being fairly inert and structurally robust, the cuticle is also permeable to small compounds and expands during growth periods between molts (1). A number of enzymes have been implicated in the shedding of old cuticles and the remodeling process that occurs as the new cuticle develops (2-8). Proteolytic enzymes have been shown to play a vital role in these processes, and inhibitor studies and rational cloning strategies have identified several nematode proteases whose functions are required for molting (9 -11).To identify novel filarial proteins involved in the molting process, we adopted a transcriptomics approach. Thousands of expressed sequence tags (ESTs) 4 have been sequenced from cDNA libraries constructed from the infective third-stage larvae (L3) and molting L3 (mL3) of the human filarial nematode Onchocerca volvulus (12, 13). Analysis of these datasets identified novel cysteine proteases involved in the molting process (14,15). Also identified in these analyses was an O. volvulus small molecular weight serine protease inhibitor (SPI) with similarities to other nematode SPI; Ascaris suum chymotrypsin/elastase in...

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Section: Discussionmentioning

confidence: 99%

Characterization of a Novel Filarial Serine Protease Inhibitor, Ov-SPI-1, from Onchocerca volvulus, with Potential Multifunctional Roles during Development of the Parasite

Ford

Guiliano

Oksov

et al. 2005

Journal of Biological Chemistry

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“…Since we have demonstrated that AceKI-1 is secreted by the adult stage of the parasite, it is possible that the hookworm releases the inhibitor in order to neutralize the potentially damaging effects of these digestive proteases within its immediate environment. In fact, other parasitic intestinal nematodes have been shown to produce potent inhibitors of chymotrypsin, pancreatic elastase, and/or trypsin, including A. suum (29,30), Trichuris suis (56), and the zoonotic nematode Anisakis simplex (57). The fact that the comparable inhibitors from A. suum and A. simplex belong to the Ascaris trypsin inhibitor family, rather than the Kunitz type, suggest that these activities have arisen in various nematodes by convergent evolution, i.e.…”

Section: Discussionmentioning

confidence: 99%

A Broad Spectrum Kunitz Type Serine Protease Inhibitor Secreted by the Hookworm Ancylostoma ceylanicum

Milstone

Harrison

Bungiro

et al. 2000

Journal of Biological Chemistry

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Although blood-feeding hookworms infect over a billion people worldwide, little is known about the molecular mechanisms through which these parasitic nematodes cause gastrointestinal hemorrhage and iron deficiency anemia. A cDNA corresponding to a secreted Kunitz type serine protease inhibitor has been cloned from adult Ancylostoma ceylanicum hookworm RNA. The translated sequence of the A. ceylanicum Kunitz type inhibitor 1 (AceKI-1) cDNA predicts a 16-amino acid secretory signal sequence, followed by a 68-amino acid mature protein with a molecular mass of 7889 daltons. Recombinant protein (rAceKI-1) was purified from induced lysates of Escherichia coli transformed with the rAceKI-1/pET 28a plasmid, and in vitro studies demonstrate that rAceKI-1 is a tight binding inhibitor of the serine proteases chymotrypsin, pancreatic elastase, neutrophil elastase, and trypsin. AceKI-1 inhibitory activity is present in soluble protein extracts and excretory/secretory products of adult hookworms but not the infective third stage larvae. The native AceKI-1 inhibitor has been purified to homogeneity from soluble extracts of adult A. ceylanicum using size exclusion and reverse-phase high pressure liquid chromatography. As a potent inhibitor of mammalian intestinal proteases, AceKI-1 may play a role in parasite survival and the pathogenesis of hookworm anemia.

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“…DNA was extracted from PBMCs using the phenol‐chloroform method, as previously described 37, 38. Briefly, blood was collected in acid‐citrate‐dextrose tubes and spun at 750 g for 8 min; then, the buffy coat containing white blood cells was aspirated and lysed in lysis buffer [0.32 M sucrose, 1% (v/v) Triton X‐100, 5 mM MgCl 2 · 6H 2 O, 12 mM TRIS HCl (pH 7.5)].…”

Section: Methodsmentioning

confidence: 99%

Genetic susceptibility to nasopharyngeal carcinoma within the HLA‐A locus in Taiwanese

Chen

Jin

et al. 2002

Intl Journal of Cancer

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NPC is an epithelial tumor that is highly prevalent among the southern Chinese. Numerous studies have indicated that specific HLA haplotypes and genes within the HLA complex are associated with NPC. As a first effort to localize the gene responsible for susceptibility, the HLA-A, -B, and -A2 subtypes were examined for their association to NPC. Consistent with previous reports, frequencies of HLA-A2 [OR ‫؍‬ 2.50, pc ‫؍‬ 0.020 (study population); OR ‫؍‬ 3.73, pc ‫؍‬ 0.0030 (>40 years old)] were significantly higher in patients with NPC than in healthy controls. Two-locus analysis indicated that A2 ؉ B46؉ individuals are at greater risk for NPC than A2 -B46 -individuals in both the population studied and the >40-year-old group. This, however, may be due to the close linkage of these 2 genes. Moreover, A2 ؉ B38؉ individuals were at higher risk than A2 -B38 -individuals in both the population studied and the >40-year-old group; A2 and B38 are not genetically linked. These findings suggest that B38 or B46 alone cannot confer a high risk of NPC but that, in conjunction with A2, B38 or B46 positivity greatly increases risk. None of 5 A2 subtypes identified from studied populations was significantly associated with NPC. Microsatellite marker D6S211, located 97 kb telomeric to HLA-A, was analyzed for its association with NPC. Allele 4 of D6S211 was significantly associated with NPC (OR ‫؍‬ 3.97, pc ‫؍‬ 0.0042). These results strongly support the hypothesis that genes associated with susceptibility to NPC in the HLA region are within the HLA-A locus.

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Anisakis simplex:Mutational Bursts in the Reactive Site Centers of Serine Protease Inhibitors from an Ascarid Nematode

Cited by 20 publications

References 18 publications

Characterization of a Novel Filarial Serine Protease Inhibitor, Ov-SPI-1, from Onchocerca volvulus, with Potential Multifunctional Roles during Development of the Parasite

Characterization of a Novel Filarial Serine Protease Inhibitor, Ov-SPI-1, from Onchocerca volvulus, with Potential Multifunctional Roles during Development of the Parasite

A Broad Spectrum Kunitz Type Serine Protease Inhibitor Secreted by the Hookworm Ancylostoma ceylanicum

Genetic susceptibility to nasopharyngeal carcinoma within the HLA‐A locus in Taiwanese

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