Thesis (MSc (Genetics. Plant Biotechnology)--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: Glycogen was isolated from E. coli and analysed for the amount of phosphate
present within it. It was confirmed that a significant proportion of the glucose residues
were phosphorylated at the C6 position. This glycogen phosphate was found also in
both glgb- (glycogen branching enzyme) and glgp- (glycogen phosphorylase enzyme)
mutants, demonstrating that a mechanism for phosphate incorporation that does not
involve GlgP alone, and which is capable of incorporating phosphate into linear
glucans could exist. The degree of phosphorylation depended on the amount of
phosphate present in the media, which less being incorporated in media where
phosphate was reduced. Screening for glycogen phosphorylating genes using a E.
coli genomic library in a functional expression system identified the malP gene as a
possible candidate for incorporation of the phosphate at the C6 position. There was
no difference, however, between the glycogen phosphate content of the mutant and
wild type. Efforts were made to construct a malp-/glgp- double mutant, but these were
unsuccessful.
In addition the influence of plants and human proteins on yeast glycogen metabolism
was also investigated. These proteins have been demonstrated to have an effect on
starch or glycogen in humans, plant and E. coli, but the data from this study indicated
that this was not the case in yeast. / AFRIKAANSE OPSOMMING: Glikogeen, wat geisoleer was uit E.coli was geanaliseer vir fosfaat inhoud daarin.
Daar was gevind dat `n beduidende proporsie van die glukose residue gefosforileerd
was op die C6 posisie. Hierdie gefosforileerde glikogeen was ook gevind in glg-
(glikogeen vertakkingsensieme) en glgp- (glikogeen fosforileringsensieme) mutante
wat daarop dui dat `n meganisme vir fosforilering bestaan was nie slegs aangewese
is op die aktiwiteit van GlgP nie, en om fosfaat te inkorporeer in linêre glukane. Die
graad van fosforilering was ook afhanklik van die hoeveelheid fosfaat teenwoordig in
die medium, met gevolglik minder wat geinkorporeer kan word in medium waar
fosfaat verminderd was. Seleksie-gebaseerde ondersoeking vir fosforileringsensieme
van glikogeen deur gebruik te maak van E. coli genomiese biblioteke in `n
funksionele uitdrukkingssisteem het die malP geen geidentifiseer as een van die
moontlike kandidate wat verantwoordelik kan wees vir inkorporering van fosfaat in
the C6 posisie. Daar was egter geen verskil in die fosfaat inhoud van glikogeen
tussen die wilde tipe en die mutante. Pogings wat aangewend is om `n malp-/glgpdubbel
mutant te konstrueer was onsuksesvol.
Verder is die invloed van plant en mens proteine op gis glikogeen ook bestudeer.
Vroeër is aangetoon dat hierdie proteine `n invloed op stysel en glikogeen het in
mense, plante en E. coli, maar data van hierdie studie toon aan dat dit nie die geval
in gis is nie.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/2524 |
| Date | 12 1900 |
| Creators | Nepembe, Mehafo, Ndafapawa |
| Contributors | Lloyd, J. R., Kossmann, J. M., University of Stellenbosch. Faculty of Agrisciences. Dept. of Genetics. Institute for Plant Biotechnology. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | Unknown |
| Type | Thesis |
| Rights | University of Stellenbosch |
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