Show simple item record

dc.contributor.authorLarge, Andrew
dc.date.accessioned2009-12-18T10:14:06Z
dc.date.available2009-12-18T10:14:06Z
dc.date.issued1995
dc.identifier.urihttp://hdl.handle.net/2436/88293
dc.descriptionA thesis submitted in partial fulfilment of the requirements of the University of Wolverhampton for the degree of Doctor of Philosophy
dc.description.abstractSeveral unusual hydrogen peroxide generating oxidase activities have been investigated in the alimentary tissues of terrestrial gastropod molluscs. A membrane-bound aromatic alcohol oxidase activity has been demonstrated in the digestive gland (hepatopancreas) of four families of gastropod (Arionidae, Limacidae, Helicidae and Achatinidae). Aromatic alcohol oxidase activity has only previously been reported in basidiomycete fungi where the activity is soluble, extracellular and probably involved in lignin biodegradation. The gastropod activity has been solubilised in nonionic detergents and the solubilised activity tested with a variety of potential substrates. In three species (Arlon ater, Limax flavus and Helix aspersa) cinnamyl alcohol (1 mM, cis/trans mixed isomers) was found to be the most favoured substrate. In the fourth, Achatina fulica, the unsaturated aliphatic alcohol crotyl alcohol (2-buten-l-ol, cis/trans mixed isomers) was found to be the most favoured substrate at 1mM although cinnamyl alcohol was also readily oxidised. All four species could oxidise crotyl alcohol although only the activity from A. fulica could oxidise saturated aliphatic primary alcohols. Alcohols up to C16 (n-hexadecyl) were utilised by this activity. Subcellular fractionation experiments showed aromatic alcohol oxidase activity to be localised in the endoplasmic reticulum. Aldehyde oxidase activity was considered in relation to its possible role in the further utilisation of the products of aromatic alcohol oxidase. It is a soluble cytosolic activity. Gastropod aldehyde oxidase is active with a variety of aldehydes but not with purines unlike the mammalian enzyme. Salicylaldehyde was found to be a specific inhibitor of the A. ater and L. flavus activities. D-Mannitol oxidase thus far has only been reported in gastropod molluscs. Activity from the four species mentioned above is very similar with regard to substrate preference and apparent Km values. o-Mannitol, D-arabitol and to a lesser extent sorbitol (D-glucitol) are oxidised. Subcellular fractionation of Arion ater digestive gland showed mannitol oxidase to be localised In a low density membrane distinct from mitochondria, peroxisomes and endoplasmic reticulum. When enriched by successive rate dependent and isopycnic sucrose gradient centrifugation the mannitol oxidase fractions were analysed by electron microscopy. This revealed the predominance of tubular structures typically of seven tubules (each of approx. 50nm in diameter) in a hexagonal 6+1 arrangement surrounded by a smooth membrane. An interim term "mannosome" has been proposed for these structures. S. D. S. -P. A. G. E. analysis revealed a single predominant polypeptide of 68-69kDa (the published value for partially purified mannitol oxidase) in the mannitol oxidase-enriched fractions. Analytical sucrose gradient centrifugation techniques provided evidence for the presence of "mannosomes" in Helix aspersa although the situation is less clear for Achatina fulica and Limax flavus.
dc.formatapplication/pdf
dc.language.isoen
dc.publisherUniversity of Wolverhampton
dc.titleAromatic alcohol oxidase, aldehyde oxidase and mannitol oxidase in terrestrial gastropods
dc.typeThesis or dissertation
dc.type.qualificationnamePhD
dc.type.qualificationlevelDoctoral
rioxxterms.licenseref.urihttps://creativecommons.org/licenses/by-nc-nd/4.0/
refterms.dateFOA2020-04-29T16:57:37Z
html.description.abstractSeveral unusual hydrogen peroxide generating oxidase activities have been investigated in the alimentary tissues of terrestrial gastropod molluscs. A membrane-bound aromatic alcohol oxidase activity has been demonstrated in the digestive gland (hepatopancreas) of four families of gastropod (Arionidae, Limacidae, Helicidae and Achatinidae). Aromatic alcohol oxidase activity has only previously been reported in basidiomycete fungi where the activity is soluble, extracellular and probably involved in lignin biodegradation. The gastropod activity has been solubilised in nonionic detergents and the solubilised activity tested with a variety of potential substrates. In three species (Arlon ater, Limax flavus and Helix aspersa) cinnamyl alcohol (1 mM, cis/trans mixed isomers) was found to be the most favoured substrate. In the fourth, Achatina fulica, the unsaturated aliphatic alcohol crotyl alcohol (2-buten-l-ol, cis/trans mixed isomers) was found to be the most favoured substrate at 1mM although cinnamyl alcohol was also readily oxidised. All four species could oxidise crotyl alcohol although only the activity from A. fulica could oxidise saturated aliphatic primary alcohols. Alcohols up to C16 (n-hexadecyl) were utilised by this activity. Subcellular fractionation experiments showed aromatic alcohol oxidase activity to be localised in the endoplasmic reticulum. Aldehyde oxidase activity was considered in relation to its possible role in the further utilisation of the products of aromatic alcohol oxidase. It is a soluble cytosolic activity. Gastropod aldehyde oxidase is active with a variety of aldehydes but not with purines unlike the mammalian enzyme. Salicylaldehyde was found to be a specific inhibitor of the A. ater and L. flavus activities. D-Mannitol oxidase thus far has only been reported in gastropod molluscs. Activity from the four species mentioned above is very similar with regard to substrate preference and apparent Km values. o-Mannitol, D-arabitol and to a lesser extent sorbitol (D-glucitol) are oxidised. Subcellular fractionation of Arion ater digestive gland showed mannitol oxidase to be localised In a low density membrane distinct from mitochondria, peroxisomes and endoplasmic reticulum. When enriched by successive rate dependent and isopycnic sucrose gradient centrifugation the mannitol oxidase fractions were analysed by electron microscopy. This revealed the predominance of tubular structures typically of seven tubules (each of approx. 50nm in diameter) in a hexagonal 6+1 arrangement surrounded by a smooth membrane. An interim term "mannosome" has been proposed for these structures. S. D. S. -P. A. G. E. analysis revealed a single predominant polypeptide of 68-69kDa (the published value for partially purified mannitol oxidase) in the mannitol oxidase-enriched fractions. Analytical sucrose gradient centrifugation techniques provided evidence for the presence of "mannosomes" in Helix aspersa although the situation is less clear for Achatina fulica and Limax flavus.


Files in this item

Thumbnail
Name:
Large_PhDthesis.pdf
Size:
10.47Mb
Format:
PDF

This item appears in the following Collection(s)

Show simple item record

https://creativecommons.org/licenses/by-nc-nd/4.0/
Except where otherwise noted, this item's license is described as https://creativecommons.org/licenses/by-nc-nd/4.0/