Parasitology Research

, Volume 80, Issue 8, pp 680–683 | Cite as

A cysteine proteinase in the cercariae ofDiplostomum pseudospathaceum (Trematoda, Diplostomatidae)

  • T. Moczoń
Original Paper

Abstract

A cysteine proteinase was detected in extracts from cercariae of the trematodeDiplostomum pseudospathaceum. The enzyme preferred protein substrates over synthetic, chromogenic peptides. The optimal pH for hydrolysis of substrates was 7.2 for azocoll, 6.4 and 7.6 for azocasein, 7.6 for azoalbumin, and 6.8 forN-benzoyl-l-arginine-4-nitroanilide. Elastin-Congo red and certainN-blockedl-aminoacyl-andl-peptidyl nitroanilides bearingl-phenylalanine,l-alanine,l-tyrosine, andl-leucine at the P1 subsite were not hydrolyzed. Thiol-reducing and divalent cation-complexing agents stimulated the proteinase activity, whereas thiol-blocking agents inhibited it. The relative molecular weight of the enzyme was approximately 40 000 as determined by SDS-PAGE. Detection of an identical proteinase in water after treatment of living cercariae with praziquantel suggests that the enzyme occupied the penetration glands in the larvae. Thus, when secreted by the parasite during invasion of an appropriate host, the enzyme might act as a penetration-promoting factor.

Keywords

Peptide Molecular Weight Cysteine Proteinase Activity Cysteine Proteinase 

Abbreviations

SDS-PAGE

sodium dodecyl sulfatepolyacrylamide electrophoresis

BSA

bovine serum albumin

TRIS

tris(hydroxymethyl)aminomethane

EGTA

ethylene glycol-O,OO′-bis(2-aminoethyl)N,N,N′,N′-tetraacetic acid

DTE

dithioerythritol

2-ME

2-mercaptoethanol

pHMB

sodiump-hydroxymercuribenzoate

PMSF

phenylmethanesulfonyl fluoride

TPCK N

tosyl-l-phenylalanine chloromethyl ketone

LCK Nα

tosyl-l-lysine chloromethyl ketone

SBTI

soybean trypsin inhibitor

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References

  1. Amiri P, Sakanari J, Basch P, Newport G, McKerrow JH (1988) TheSchistosomatium douthitti cercarial elastase is biochemically and structurally distinct from that ofSchistosoma mansoni. Parasitol Res 77: 697–702Google Scholar
  2. Baba EH, Homewood CA, Gazzinelli G, Atkinson EM (1977) Comparison of proteolytic enzymes from cercarial extract and secretion. Comp Biochem Physiol [B] 57: 55–57Google Scholar
  3. Charney J, Tomarelli RM (1947) A colorimetric method for the determination of the proteolytic activity of duodenal juice. J Biol Chem 171: 501–505Google Scholar
  4. Gazzinelli G, Ramalho-Pinto FJ, Pellegrino J (1966) Purification and characterization of the proteolytic enzyme complex of cercarial extract. Comp Biochem Physiol [B] 18: 689–700Google Scholar
  5. Gazzinelli G, Mares-Guia M, Pellegrino J (1972) Reaction of the main fraction ofSchistosoma mansoni cercarial enzymes with synthetic substrates and inhibitors of proteolytic enzymes. Exp Parasitol 32: 21–25PubMedGoogle Scholar
  6. Hjertén S (1962) “Molecular sieve” chromatography on polyacrylamide gels, prepared according to a simplified method. Arch Biochem Biophys [Suppl 1]: 147–151PubMedGoogle Scholar
  7. Klein IB, Kirsch JF (1969) The activation of papain and the inhibition of the active enzyme by carbonyl reagents. J Biol Chem 244: 5928–5935PubMedGoogle Scholar
  8. Laemmli UK (1970) Cleavage of structural proteins during the assembly of bacteriophage T4. Nature 227: 680–685PubMedGoogle Scholar
  9. Matsumura K, Shimada M, Sato K, Aoki Y (1990) Praziquantelinduced secretion of proteolytic enzyme fromSchistosoma mansoni cercariae. J Parasitol 76: 436–438PubMedGoogle Scholar
  10. McKerrow JH, Pino-Heiss S, Lindquist R, Werb Z (1985) Purification and characterization of an elastinolytic proteinase secreted by cercariae ofSchistosoma mansoni. J Biol Chem 260: 3703–3707PubMedGoogle Scholar
  11. Peterson GL (1983) Determination of total protein. Methods Enzymol 91: 95–119PubMedGoogle Scholar
  12. Reisfeld R, Lewis U, Williams D (1962) Disc electrophoresis of basic proteins and peptides on polyacrylamide gels. Nature 195: 281–283PubMedGoogle Scholar
  13. Schechter I, Berger A (1967) On the size of the active site in proteases. I. Papain. Biochem Biophys Res Commun 27: 157–162PubMedGoogle Scholar
  14. Schotton DM (1970) Elastase. Methods Enzymol 19: 113–140Google Scholar
  15. Tomarelli RM, Charney J, Harding ML (1949) The use of azoalbumin as a substrate in the colorimetric determination of peptic and tryptic activity. J Lab Clin Med 34: 428–433Google Scholar

Copyright information

© Springer-Verlag 1994

Authors and Affiliations

  • T. Moczoń
    • 1
  1. 1.Institute of ParasitologyPolish Academy of SciencesWarsawPoland

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