A new form of arthropod phenoloxidase is abundant in venom of the parasitoid wasp Pimpla hypochondriaca

Parkinson, Neil; Smith, Ian R.; Weaver, Robert J.; Edwards, John

doi:10.1016/s0965-1748(00)00105-3

Cited by 93 publications

(74 citation statements)

References 32 publications

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“…Various fungi, several insects (Thomas et al, 1989), bacteria (Alexandre & Zhulin, 2000 ;Sanchez-Amat et al, 2001 ;Martins et al, 2002) and recently wasp venom (Parkinson et al, 2001) have been shown to produce\ contain laccase. The biological role for laccase has as yet not been fully elucidated and appears to vary depending on the type of organism (Thurston, 1994).…”

Section: Introductionmentioning

confidence: 99%

Molecular cloning of a laccase isozyme gene from Pleurotus sajor-caju and expression in the heterologous Pichia pastoris host The GenBank accession number for the sequence of the Pleurotus sajor-caju laccase 4 gene (lac4) reported in this paper is AF297228.

2002

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The Psc lac4 gene from Pleurotus sajor-caju has been cloned and expressed in the heterologous host Pichia pastoris, under the control of the AOX1 methanol inducible promoter. The native Ple. sajor-caju laccase signal sequence was effective in directing the secretion of lac4 expressed in Pic. pastoris. The control of media pH and temperature was found to be important in obtaining sufficient quantities of the protein to allow purification and subsequent biochemical characterization. The recombinant Psc Lac4 was purified to electrophoretic homogeneity and was shown to be immunologically related to Pleurotus eryngii Lac1. The purified laccase was estimated to have a molecular mass of around 59 kDa, to have a carbohydrate content of approximately 7 % and a calculated pI of 438. The enzyme oxidized the substrates 2,2-azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS), 2,6-dimethoxyphenol, syringaldazine and guaiacol, exhibiting optimal pHs of 33, 6, 65 and 7 respectively. With ABTS as substrate the enzyme displayed optimal activity at 35 mC and pH 35. The enzyme was strongly inhibited by sodium azide and thioglycolic acid but not by EDTA.

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Section: Introductionmentioning

confidence: 99%

Molecular cloning of a laccase isozyme gene from Pleurotus sajor-caju and expression in the heterologous Pichia pastoris host The GenBank accession number for the sequence of the Pleurotus sajor-caju laccase 4 gene (lac4) reported in this paper is AF297228.

2002

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“…Encapsulation and melanization of invading foreign organisms in insects are associated with an enzyme cascade in which the terminal component is tyrosinase (11-13). Loss of blood in insects during wounding is prevented by melanin deposition at the damaged site (14). Tyrosinase in association with other enzymes generates reactive intermediates leading to cross-linking of structural proteins and chitin to form hardened cuticle, which is a vital process for the survival of insects and anthropods (15-18).…”

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confidence: 99%

Oxyresveratrol and Hydroxystilbene Compounds

Kim

Yun

Lee

et al. 2002

Journal of Biological Chemistry

285

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Tyrosinase is responsible for the molting process in insects, undesirable browning of fruits and vegetables, and coloring of skin, hair, and eyes in animals. To clarify the mechanism of the depigmenting property of hydroxystilbene compounds, inhibitory actions of oxyresveratrol and its analogs on tyrosinases from mushroom and murine melanoma B-16 have been elucidated in this study. Oxyresveratrol showed potent inhibitory effect with an IC 50 value of 1.2 M on mushroom tyrosinase activity, which was 32-fold stronger inhibition than kojic acid, a depigmenting agent used as the cosmetic material with skin-whitening effect and the medical agent for hyperpigmentation disorders. Hydroxystilbene compounds of resveratrol, 3,5-dihydroxy-4 -methoxystilbene, and rhapontigenin also showed more than 50% inhibition at 100 M on mushroom tyrosinase activity, but other methylated or glycosylated hydroxystilbenes of 3,4 -dimethoxy-5-hydroxystilbene, trimethylresveratrol, piceid, and rhaponticin did not inhibit significantly. None of the hydroxystilbene compounds except oxyresveratrol exhibited more than 50% inhibition at 100 M on L-tyrosine oxidation by murine tyrosinase activity; oxyresveratrol showed an IC 50 value of 52.7 M on the enzyme activity. The kinetics and mechanism for inhibition of mushroom tyrosinase exhibited the reversibility of oxyresveratrol as a noncompetitive inhibitor with L-tyrosine as the substrate. The interaction between oxyresveratrol and tyrosinase exhibited a high affinity reflected in a K i value of 3.2-4.2 ؋ 10 ؊7 M. Oxyresveratrol did not affect the promoter activity of the tyrosinase gene in murine melanoma B-16 at 10 and 100 M. Therefore, the depigmenting effect of oxyresveratrol works through reversible inhibition of tyrosinase activity rather than suppression of the expression and synthesis of the enzyme. The number and position of hydroxy substituents seem to play an important role in the inhibitory effects of hydroxystilbene compounds on tyrosinase activity.Tyrosinase catalyzes two distinct reactions of the conversion of tyrosine to Dopa 1 (tyrosine hydroxylase activity (EC 1.14.18.1); tyrosine, 3,4-dihydroxyphenylalanine, oxygen, and oxidoreductase) and the oxidation of the resultant Dopa (Dopa oxidase activity (EC 1.10.3.1); Dopa, oxygen, and oxidoreductase) (1, 2). The enzyme oxidizes phenols and diphenols using a catalytic mechanism that depends on the presence of copper atoms at the active site (3, 4). Dopaquinone produced by tyrosinase is nonenzymatically converted to dopachrome, which is acted upon by an isomerase producing dihydroxyindoles (5, 6). Melanin pigments are eventually produced by further oxidation and polymerization of the indoles (7).Tyrosinase, an enzyme catalyzing the rate-limiting step for the biosynthetic pathway of melanin pigments, is widely distributed in nature. It exists in many organisms with slightly different forms. The enzyme participates in several important reactions of host defense, wound healing, and sclerotization in insects and other arthropods (8 -10). E...

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“…When injected into host larvae, it was capable of inducing developmental arrest that resembled natural parasitism (Coudron and Brandt, 1996). Molecular and biochemical analyses of two wasp species (Pimpla hypochondriaca and P. turionella) from the genus Pimpla suggest that these endoparasitoid venoms possess a large and diverse number of enzymes, including laccase, serine protease, reprolysin-like metalloprotease, phospholipases, and phenoloxidases (Parkinson et al, 2001(Parkinson et al, , 2002a(Parkinson et al, ,b, 2003Uçkan et al, 2004). Uçkan et al (2004) also observed the presence of noradrenaline, apamin, and melittin in venom from P. turionella, consistent with the paralytic action of the venom in multiple life stages of lepidopteran hosts (Kansu and Uur, 1984).…”

Section: January 2006mentioning

confidence: 99%

“…To confirm the presence of these agents, venom assays were performed with pure apamin (Sigma) and anti-histamine antibodies (Sigma), either alone or in combination with a LC 99 dose of crude venom from N. vitripennis (Rivers et al, 1993). Similarly, phenoloxidases have been identified as major venom components of some wasp species (Dani et al, 2003;Parkinson et al, 2001), as well as components of parasitoid larval secretions, including N. vitripennis (Gerling and Legner, 1968;Thompson, 1986;Whiting, 1967). Consequently, parallel experiments were conducted with anti-phenoloxidase antibodies (Research Diagnostics, Flanders, NJ) either alone or simultaneously with crude venom.…”

Section: Venom Assaysmentioning

confidence: 99%

“…The infrared spectrum also revealed the presence of phosphorus-containing structures, typical of enzymes, in the venom. Though the presence of enzymatic activity in N. vitripennis venom has yet to be confirmed, venoms from several endoparasitic species and social Hymenoptera have been shown to contain multiple types of enzymes (Moreau et al, 2004;Parkinson et al, 2001;2002a,b;Piek and Spanjier, 1986;Schmidt, 1982;Uçkan et al, 2004). Enzyme activity, however, does not likely account for the ability of venom from N. vitripennis to arrest host development or evoke death as crude venom preparations contained the protease inhibitor PMSF as well as the chelating agent EDTA, and high concentrations of anti-phenoloxidase antibodies had no influence on venom activity.…”

Section: January 2006mentioning

confidence: 99%

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Characterization and biochemical analyses of venom from the ectoparasitic waspNasonia vitripennis (Walker) (hymenoptera: Pteromalidae)

Rivers

Uçkan

Ergın

2005

Arch. Insect Biochem. Physiol.

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During parasitism, the ectoparasitic wasp Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae) induces a developmental arrest in host pupae that is sustained until the fly is either consumed by developing larvae or the onset of death. Bioassays using fluids collected from the female reproductive system (calyx, alkaline gland, acid gland, and venom reservoir) indicated that the venom gland and venom reservoir are the sources of the arrestant and inducer(s) of death. Infrared spectroscopic analyses revealed that crude venom is acidic and composed of amines, peptides, and proteins, which apparently are not glycosylated. Reversed phase high performance liquid chromatography (HPLC) and sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) confirmed the proteinaceous nature of venom and that it is composed mostly of mid to high molecular weight proteins in the range of 13 to 200.5 kilodaltons (kDa). Ammonium sulfate precipitation and centrifugal size exclusion membranes were used to isolate venom proteins. SDS-PAGE protein profiles of the isolated venom fractions displaying biological activity suggest that multiple proteins contribute to arresting host development and eliciting death. Additionally, HPLC fractionation coupled with use of several internal standards implied that two of the low molecular weight proteins were apamin and histamine. However, in vitro assays using BTI-TN-5B1-4 cells contradict the presence of these agents. Abbreviations used: HPLC = high performance liquid chromatography; I.R. = infrared spectroscopy; kDa = kilodalton; LC 99 = lethal concentration to kill 100% of population; LD = light-dark; MWCO = molecular weight cutoff; SDS-PAGE = sodium dodecyl sulfate polyacrylamide gel electrophoresis; TFA = trifluoroacetic acid; VRE = venom reservoir equivalent.

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A new form of arthropod phenoloxidase is abundant in venom of the parasitoid wasp Pimpla hypochondriaca

Cited by 93 publications

References 32 publications

Molecular cloning of a laccase isozyme gene from Pleurotus sajor-caju and expression in the heterologous Pichia pastoris host The GenBank accession number for the sequence of the Pleurotus sajor-caju laccase 4 gene (lac4) reported in this paper is AF297228.

Molecular cloning of a laccase isozyme gene from Pleurotus sajor-caju and expression in the heterologous Pichia pastoris host The GenBank accession number for the sequence of the Pleurotus sajor-caju laccase 4 gene (lac4) reported in this paper is AF297228.

Oxyresveratrol and Hydroxystilbene Compounds

Characterization and biochemical analyses of venom from the ectoparasitic waspNasonia vitripennis (Walker) (hymenoptera: Pteromalidae)

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