Life as we know it would be impossible in the absence of oxygen. However,
too much oxygen can be toxic to the aerobic organisms which depend on it
for their very existence. This apparent paradox arises as a result of oxygen's
ability to accept electrons, forming highly reactive (reduced) oxygen species
such as superoxide, hydrogen peroxide and the hydroxyl radical. The toxicity
of oxygen is greatly enhanced in illuminated plants, due to the photosynthetic
reactions which produce both oxygen and highly energetic electrons in close
proximity to one another. These problems are further exacerbated when
plants are exposed to a variety of stress conditions, since these conditions
reduce the ability of plants to utilise excess electrons. As a result of the
danger posed by· these reactive oxygen species, plants have· evolved a
complex antioxidant system for their scavenging.
Research has shown that plants with naturally elevated levels of the
components of the antioxidant system are better equipped to deal with stress
conditions which enhance the production of reactive oxygen species. A
considerable amount of research has thus been dedicated to the elucidation
of the antioxidant system. Almost as much research has been dedicated to
enhancing the antioxidant system, with the aim of improving plant productivity
under stress conditions.
This study sought to evaluate plants carrying elevated levels of two of the
enzymes of the antioxidant system. For these purposes, tobacco was
transformed with the gene for E. coli glutathione reductase (GR), an enzyme
believed to catalyse the rate limiting reaction in the scavenging of hydrogen
peroxide. This gene was fused to the gene for the RUBISCO small subunit
transit peptide - a peptide capable of targeting proteins to the chloroplast.
Due to the presence of this peptide the transformed plants exhibited high
chloroplastic levels of GR activity. These plants were crossed with a second
tobacco transformant carrying high levels of chloroplastic tomato superoxide
dismutase (SOD) - an enzyme responsible for the scavenging of superoxide.
These hybrid plants were shown to exhibit high GR and SOD activities in the
chloroplast .- the subcellular compartment most susceptible to damage
caused by reactive oxygen species.
The transgenic hybrids were evaluated for their ability to tolerate oxidative
stress by treating them with paraquat - a herbicide whose mode of action
involves the production of large quantities of activated oxygen. Under stress
conditions, plants carrying just E. coli GR showed a slight improvement in
their ability to deal with oxidative stress. In contrast to this, the SOD
transformants showed more cellular damage than untransformed control
plants. This was attributed to the inability of other enzymes in the antioxidant
pathway to deal with the increased flow of metabolites through the pathway.
The hybrid transformants showed enhanced stress tolerance in the initial
stages of oxidative stress, but this declined with ongoing exposure to stress
conditions. As with the SOD transformants, this decline in protection was
. ascribed to the relatively low activities of the other enzymes in the antioxidant pathway. It was concluded that elevated levels of the two enzymes conferred
greater stress tolerance than just one of the enzymes, but for true stress
tolerance it will be necessary to evaluate the antioxidant system and enhance
the activity of further enzymes in the pathway. It may also be necessary to
improve the regulation of transgene expression, ensuring that none of the
enzymes are overwhelmed by the increased flow of metabolites through the
system. / Thesis (M.Sc.)-University of Natal, 1996
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ukzn/oai:http://researchspace.ukzn.ac.za:10413/4928 |
| Date | January 1996 |
| Creators | Penter, Mark Gavin. |
| Contributors | Amory, Alan M. |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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