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Current Research
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Engineering Plant Phytochelatin Biosynthesis:
Towards Phytoremediation
Harnessing the power of nature for environmental cleanup
offers a promising biotechnology for improving global
health. Heavy metal contamination poses health and
environmental challenges with a price tag for remediation
estimated at $200 billion in this country alone. Since the
costs of growing and harvesting a crop are minimal compared
to those of soil replacement, the use of plants for
remediation of hazardous soil shows potential; however, no
single plant currently displays all the necessary traits for
efficient phytoremediation. Therefore, understanding the
molecular basis for how plants protect themselves from metal
toxicity and engineering these processes are crucial for
optimizing plants as agents for phytoremediation.
Phytochelatin peptides (Figure 2) play a role in heavy metal
tolerance by chelating metal ions for sequestration in
vacuoles. Derived from glutathione by the action of
phytochelatin synthase (PCS), phytochelatins consist of
repeating units of y-glutamylcysteine. This project employs
a strategy directed at elucidating the molecular basis for
phytochelatin production with the goal of re-engineering PCS
for improved heavy metal protection properties. In
collaboration with Phil Rea (U. Penn), studies of PCS aim to
understand its reaction mechanism and substrate/product
specificity. With this knowledge, protein engineering of may
lead to the creation of a molecular system for plant-based
detoxification of heavy metal contaminated soils. Click here to read a St. Louis Post-Dispatch Article about this research.
Plant Glutathione Biosynthesis: A General Stress
Response
Regulation of the intracellular redox environment is
critical in cellular physiology for influencing signaling
pathways and cell fate in response to stress. In plants, the
peptide glutathione plays multiple roles as protection
against various environmental stresses. As an antioxidant,
glutathione quenches reactive oxygen species that cause
cellular damage. Plants also use glutathione for the
detoxification of xenobiotics, herbicides, air pollutants
such as sulfur dioxide and ozone, and heavy metals. For
example, exposure to heavy metals increases the production
of glutathione as a general protection mechanism and
provides metabolic precursors for formation of the heavy
metal chelating phytochelatin molecules.
Although glutathione accumulates in response to different
stress stimuli in plants, the structural and functional
properties of the plant enzymes responsible for its
production remain biochemically uncharacterized. We are
currently studying the two enzymes, glutamate-cysteine
ligase and glutathione synthetase, that catalyze glutathione
biosynthesis in plants. Understanding how these enzymes
catalyze the biosynthesis of glutathione and how their
enzymatic activities are modulated will provide a
fundamental understanding of how these proteins function to
protect plants from environmental stresses.
Cysteine Synthase Complex: A Macromolecular
Sensor Regulating Sulfur Assimilation
Sulfur is an essential nutrient for plant growth and
development. In plant sulfur assimilation, the biosynthesis
of the amino acid cysteine has a central role in controlling
the availability of reduced sulfur to the plant and in
providing cysteine as a precursor for all cellular compounds
containing sulfur. Sulfur deficiency is a growing problem
for agriculture that results in decreased crop quality and
yields. Although studies on the mechanisms for controlling
sulfur uptake and assimilation suggest approaches for
manipulating gene expression to produce crops with improved
sulfur utilization and sulfur-deficiency stress tolerance,
the lack of basic knowledge about the underlying metabolic
regulatory network currently hampers the design of
transgenic crops with improved sulfur metabolism.
This project focuses on two enzymes that catalyze the
chemical reactions leading to cysteine formation -- serine
acetyltransferase (SAT) and O-acetylserine sulfhydrylase (OASS).
A key feature of this pathway is the physical association of
SAT and OASS to form the cysteine synthase (CS) complex
(Figure 3). This multi-enzyme complex acts as a molecular
sensor in the regulatory circuit that coordinates sulfate
assimilation and modulates intracellular cysteine levels. We
are seeking to establish a structural model for how the CS
complex functions as a sensor in this network.
Phosphatidylcholine Biosynthesis in C. elegans: A
New Drug Target?
Parasitic nematodes are major causes of human, animal, and
plant diseases worldwide. Although a number of therapeutics
are available as treatments, reported resistance to certain
anthelmintics, severe side-effects, or limited efficacy
resulting from differences in the life cycles of target
organisms underscore the need for the continued development
of nematocidal compounds. Identifying biochemical targets
that differ between the parasite and host species is
essential for finding effective new molecules. The
free-living nematode Caenorhabditis elegans serves as a
useful model system for studying nematode biology and for
analyzing the biochemistry of enzymes in potential target
pathways. Providing a major component of cellular membranes,
the core metabolic pathways of phosphatidylcholine synthesis
in eukaryotes are well conserved; however, recent studies
suggest that nematodes (and Plasmodia) use a different
metabolic route to this phospholipid than mammals. In
addition, phosphatidylcholine is a precursor in the
production of glycoconjugates secreted by parasitic
nematodes to avoid host immune responses. We are
collaborating with Divergence, Inc. (www.divergence.com) to
mechanistically and structurally characterize new protein
targets for the development of compounds targeting parasitic
nematodes.
Figure 2. Chemical structure of a phytochelatin. The core
structure consists of multiple y-glutamylcysteines derived
from glutathione; the one shown is condensed from two
glutathione molecules.
Figure 3. Interaction of OASS with the C-terminus of SAT.
View of the molecular surface of Arabidopsis OASS forming
the binding site. The C-terminal Arabidopsis SAT peptide
(rose) is shown as a stick drawing. The surface
corresponding to the active site is colored yellow. The
surface of residues previously implicated in interaction
between OASS and SAT are shown in green. |
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