Advertisement

Protoplasma

, Volume 175, Issue 3–4, pp 93–101 | Cite as

The association of mannitol oxidase with a distinct organelle in the digestive gland of the terrestrial slugArion ater

  • A. T. Large
  • Carolyn J. P. Jones
  • M. J. Connock
Article

Summary

Post nuclear supernatants prepared from homogenates of the digestive gland of the slugArion ater were analysed by rate-dependent and density-dependent density gradient zonal centrifugation. Mannitol oxidase carrying structures exhibited distinct centrifugal behaviour sedimenting more slowly than mitochondria, lysosomes and peroxisomes but faster than microsomes, and banding sharply at a density of 1.15 g/ml in sucrose gradients. By combining rate and isopycnic centrifugations mannitol oxidase carrying structures were largely separated from contaminating organelles. Electron microscopic examination of such mannitol oxidase-enriched fractions revealed the predominant presence of distinct structures of tubular form. SDS PAGE analysis indicated that the major polypeptide present had a mass of ∼ 68 kDa corresponding to the major subunit previously reported for partially purified mannitol oxidase ofHelix aspersa.

Keywords

Subcellular fractionation Arion ater Digestive gland Mannitol oxidase Novel organelle 

Abbreviations

ER

endoplasmic reticulum

SDS PAGE

sodium dodecylsulphate polyacrylamide gel electrophoresis

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Babson AL, Greeley SJ, Coleman CM, Phillips GE (1966) Phenolphthalein monophosphate as a substrate for serum alkaline phosphatase. Clin Chem 12: 482–490PubMedGoogle Scholar
  2. Baum H, Dodgson KS, Spencer B (1959) The assay of aryl sulphatase A and B in human urine. Clin Chem Acta 4: 453–455Google Scholar
  3. Beaufay H, Amar-Costesec A, Feytmans E, Thines Sempoux D, Wibo K, Robbi M, Berthet J (1974) Analytical study of the microsomes and isolated subcellular membranes from rat liver I. Biochemical methods. J Cell Biol 61: 188–200Google Scholar
  4. Cornejo CR (1987) Modificaciones ultrastructurales del epitelio del tubo disgestivo anterior en ejemplares en ayunas deTheba pisana (Mollusca: Gastropoda: Pulmonata). Iberus 7: 153–164Google Scholar
  5. De Duve C (1967) General principles. In: Roodyn DB (ed) Enzyme cytology. Academic Press, New York, pp 1–26Google Scholar
  6. Fishman WH, Stahl PD (1984) β-D-glucuronidase. In: Bergmyer HU (ed) Methods in enzymatic analysis, vol 4. Academic Press, New York, pp 246–256Google Scholar
  7. Gorgas K, Völkl A (1984) Peroxisomes in sebaceous glands. IV Aggregates of tubular peroxisomes in mouse Meibomian gland. Histochemistry 16: 1079–1098Google Scholar
  8. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685PubMedGoogle Scholar
  9. Lanzetta PA, Alvarez LJ, Reinach PS, Candia OA (1979) An improved assay for nanomole amounts of inorganic phosphate. Anal Biochem 100: 95–97PubMedGoogle Scholar
  10. Large AT, Connock MJ (1993) Aromatic alcohol oxidase and aldehyde oxidase activities in the digestive gland of three species of terrestrial gastropod. Comp Biochem Physiol 104 B: 489–491Google Scholar
  11. Malik ZM, Jones CJP, Connock MJ (1987) Assay and subcellular localization of H2O2-generating mannitol oxidase in the terrestrial slugArion ater. J Exp Zool 242: 9–15Google Scholar
  12. Mann V, Large AT, Khan S, Malik Z, Connock MJ (1989) Aromatic alcohol oxidase: a new membrane-bound H2O2-generating enzyme in alimentary tissues of the slugArion ater. J Exp Zool 251: 265–274Google Scholar
  13. Moya J, Rallo AM (1975) Intracisternal polycylinders: a cytoplasmic structure in cells of the terrestrial slugArion empiricorum Férussac (Pulmonata, Stylommatophora). Cell Tissue Res 159: 423–433Google Scholar
  14. Vorhaben JE, Scott JF, Campbell JW (1980) D-mannitol oxidation in the land snailHelix aspersa. J Biol Chem 255: 1950–1955PubMedGoogle Scholar
  15. —, Smith DD, Campbell JW (1984) Histochemical and biochemical localization of mannitol oxidase in the snailHelix aspersa. J Exp Zool 231: 157–160Google Scholar
  16. — — — (1986) Mannitol oxidase: partial purification and characterisation of the membrane-bound enzyme from the snailHelix aspersa. Int J Biochem 18: 337–344PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • A. T. Large
    • 1
  • Carolyn J. P. Jones
    • 2
  • M. J. Connock
    • 1
  1. 1.School of Applied SciencesUniversity of WolverhamptonWolverhamptonEngland
  2. 2.Department of Pathological Sciences, Medical SchoolUniversity of ManchesterManchester

Personalised recommendations