Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.238856
Title: Investigation of the basis of specific secretion of cellulase CelV by the Out apparatus of Erwinia carotovora subspecies carotovora
Author: Walker, Denise S.
ISNI:       0000 0001 3553 0544
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 1994
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Abstract:
Erwinia species cause soft rot in a range of economically important plants. The secretion of cellulases and pectinases, via the Out apparatus is considered a major pathogenic determinant. The Out proteins are encoded by a cluster of at least 13 genes, outC to outO, and show homology to the pullulanase secretion proteins of Klebsiella oxytoca, and secretion proteins of other plant and animal pathogens. This study concerns the nature of the interaction between the Out apparatus and the secreted enzymes, with a view to identification of the recognition motifs; on the secreted proteins that allow specific secretion. Secretion deficient mutants were isolated using hydroxylamine and PCR mutagenesis. These were characterised with respect to activity and stability of the Ce1V derivatives; ability to secrete other proteins; and DNA sequence. Two classes of mutant were identified. The first class exhibited accumulation of a cata1ytica11y active but conformationally defective product which did not affect the secretion of other proteins. These were therefore predicted to be defective at the level of recognition. The second class exhibited accumulation of a stable, full size product which interfered with the secretion of pectate lyase and were therefore predicted to be blocked within the translocation apparatus. The isolation of intragenic suppressors of some of these mutants is described. Several deleted derivatives of Ce1V were constructed, all of which were severely affected in their ability to be secreted, even when only-12 carboxy terminal amino acids were affected. In conclusion, recognition for secretion is highly dependent on conformational integrity of both of the functional domains of which the enzyme is composed. The pathways followed by cellulase and pectinases appear to be initially separate, converging to a common translocation apparatus. In addition, cellulase, although having limited independent macerating ability, was identified as playing a vital role in the collaborative action of exoenzymes during attack of potato tubers.
Supervisor: Not available Sponsor: Natural Environment Research Council ; British Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.238856  DOI: Not available
Keywords: QK Botany
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