Bicz, Joanna (2013) Investigations of siderophore and tetronic acid biosynthesis in streptomyces scabies 87.22. PhD thesis, University of Warwick.

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Abstract

Streptomyces are Gram-positive bacteria, usually found living within the soil and
they are saphrophytes. Among this class of bacteria are some plant pathogenic
species, which cause infection of the roots or the tubers of some plants. The model
Streptomycete plant pathogen is Streptomyces scabies; this infects root crops, such as
potato or radish and is a known cause of scab disease. Most Streptomyces species are
producers of secondary metabolites, many of which possess important biological
activities, such as antibacterial, iron-chelating, anticancer or immunosuppressant.
One group of these secondary metabolites are called siderophores. These are small
organic molecules, which can chelate ferric iron. The iron in the environment is
mainly present as iron (III) hydroxide, which is not very water soluble and cannot,
therefore, be taken up directly by microorganisms. Some bacteria solve this problem
through production of siderophores. The siderophores are released into the
environment by the microorganisms to chelate iron (III) from the environment and
transport it into the cell across the cell membrane. Iron is required for many life
processes.
Analysis of the Streptomyces scabies genome sequence resulted in the identification
of gene clusters predicted to direct the biosynthesis of known siderophores, e.g.
desferrioxamines and pyochelin, as well as, potentially novel siderophores. A gene
inactivation and comparative metabolic profiling approach has been employed to
identify the metabolic products of these gene clusters.
A PCR-targeting method was used to replace part of or whole genes in the S. scabies
87.22 putative secondary metabolite gene clusters. An internal fragment of the
scabichelin biosynthetic gene scab85471 and the putative S. scabies desC gene were
deleted using this method. The scabichelin and desC gene mutants were subsequently
analysed by LC-MS allowing confirmation of the function of the genes investigated.
Production of scabichelin by S. scabies 87.22 wild type was analysed by comparing
it with the authentic standard. The chemical and genetic complementation of the Δ
desC mutant was carried out to establish the involvement of the desC gene in the
biosynthesis of desferrioxamines.
The S. scabies 87.22 cryptic tetronate biosynthetic gene cluster predicted to encode a
novel agglomerin-like product, which could potentially be involved in plant
pathogenicity was also investigated. The expression of the gene cluster was first
analysed using reverse transcriptase PCR (RT-PCR) which was carried out on the
total RNA isolated from the wild type S. scabies. Following this, an attempt was
made to disrupt the scab63021 gene, a putative transcriptional activator of the cryptic
tetronate-like cluster in the S. scabies genome. Transcriptional analysis of the wild
type S. scabies and the putative Δscab63021 mutant genomes did not show any
difference in the expression of the tetronate genes between the wild type strain and
the Δ scab63021 mutant.

Item Type:	Thesis or Dissertation (PhD)
Subjects:	Q Science > QP Physiology Q Science > QR Microbiology
Library of Congress Subject Headings (LCSH):	Siderophores -- Synthesis, Streptomyces scabies, Microbial genomes, Polymerase chain reaction
Official Date:	March 2013
Dates:	Date Event March 2013 Submitted
Institution:	University of Warwick
Theses Department:	Department of Chemistry
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Challis, Gregory L.
Sponsors:	University of Warwick; United States. Department of Agriculture
Extent:	xx, 200 leaves : illustrations.
Language:	eng

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