Controlling the rate of transcription and translation of Rubisco transgenes effectively in higher-plant plastids. Genetic transformation of the circular genome of the plastids provides a containable means for modifying plant growth by manipulating photosynthesis. Although the transformation mechanism is precise, predicting the level of foreign gene expression is difficult because the amounts of messenger RNA and protein produced by foreign genes in plastids varies widely, even when the protein a ....Controlling the rate of transcription and translation of Rubisco transgenes effectively in higher-plant plastids. Genetic transformation of the circular genome of the plastids provides a containable means for modifying plant growth by manipulating photosynthesis. Although the transformation mechanism is precise, predicting the level of foreign gene expression is difficult because the amounts of messenger RNA and protein produced by foreign genes in plastids varies widely, even when the protein assembles without difficulty. This project will devise strategies for controlling this variability that will facilitate attempts to exploit plastid transformation for transplanting better versions of the photosynthetic CO2-fixing enzyme, Rubisco, into plants to improve their growth efficiency in terms of water, fertiliser and light use.Read moreRead less
Practical strategies for engineering the CO2-fixing enzyme, Rubisco, whose subunits are encoded in different subcellular compartments. My recent replacement of the plant CO2-fixing enzyme, Rubisco, with a less efficient bacterial version, with a single type of subunit encoded by a single gene, demonstrated the feasibility of replacing Rubisco. This encourages ongoing attempts to replace plant Rubisco with more efficient versions that would allow the plants to grow with less water, fertiliser or ....Practical strategies for engineering the CO2-fixing enzyme, Rubisco, whose subunits are encoded in different subcellular compartments. My recent replacement of the plant CO2-fixing enzyme, Rubisco, with a less efficient bacterial version, with a single type of subunit encoded by a single gene, demonstrated the feasibility of replacing Rubisco. This encourages ongoing attempts to replace plant Rubisco with more efficient versions that would allow the plants to grow with less water, fertiliser or light. The most efficient Rubiscos are more complex, with two different types of subunits which, in plants, are encoded in different subcellular compartments (nucleus and plastid). This proposal addresses the challenges associated with complementary engineering both genomes to substitute foreign Rubiscos into higher-plant chloroplasts.Read moreRead less
Enhancing plant photosynthesis by engineering the carbon dioxide (CO2)-fixing enzyme Rubisco. Improving the ability of crops to use water, sunlight and fertiliser more efficiently would have economic benefits for Australia and ease the environmental impacts associated with agricultural practices. Photosynthesis research has confirmed that such improvements are theoretically possible by enhancing the efficiency of the protein, Rubisco, which initiates the conversion of carbon dioxide into carbon ....Enhancing plant photosynthesis by engineering the carbon dioxide (CO2)-fixing enzyme Rubisco. Improving the ability of crops to use water, sunlight and fertiliser more efficiently would have economic benefits for Australia and ease the environmental impacts associated with agricultural practices. Photosynthesis research has confirmed that such improvements are theoretically possible by enhancing the efficiency of the protein, Rubisco, which initiates the conversion of carbon dioxide into carbon compounds required for growth. The biotechnological research proposed here uses unique capabilities to improve our understanding of structural features in Rubisco that influence its assembly and functional efficiency in plants. This knowledge will pave the way for transplanting more efficient Rubisco into crops to improve their growth.Read moreRead less
Organophosphate pesticide degradation: evolved enzymes and biomimetics for bioremediation and medicine. Organophosphate (OP) pesticides are an indispensable part of modern agriculture - their use results in dramatically increased crop yields. However, they are toxic and can damage the environment and cause significant health problems. Enzymes are currently being used to treat runoff water that is contaminated with OPs. The same enzymes also have the potential to aid in the treatment of OP poison ....Organophosphate pesticide degradation: evolved enzymes and biomimetics for bioremediation and medicine. Organophosphate (OP) pesticides are an indispensable part of modern agriculture - their use results in dramatically increased crop yields. However, they are toxic and can damage the environment and cause significant health problems. Enzymes are currently being used to treat runoff water that is contaminated with OPs. The same enzymes also have the potential to aid in the treatment of OP poisoning. However, OP degrading enzymes could be improved in many ways - we will evolve these enzymes to enhance their catalytic properties - to enable them to act more efficiently on an increased number of OPs. Read moreRead less
Identifying potential barriers to transplanting modified forms of the CO2-fixing enzyme, Rubisco, into plants. Improving the ability of crops to use water, light and fertiliser more efficiently would have economic benefits and ease the environmental impacts associated with agricultural practices. It is thought that such improvements can be made by enhancing the efficiency of the photosynthetic protein, Rubisco, which fixes most of the CO2 in the biosphere. The research proposed here uses unique ....Identifying potential barriers to transplanting modified forms of the CO2-fixing enzyme, Rubisco, into plants. Improving the ability of crops to use water, light and fertiliser more efficiently would have economic benefits and ease the environmental impacts associated with agricultural practices. It is thought that such improvements can be made by enhancing the efficiency of the photosynthetic protein, Rubisco, which fixes most of the CO2 in the biosphere. The research proposed here uses unique Rubisco transplantation capabilities that I have developed to improve our fundamental understanding of how Rubisco is processed and its activity regulated in plants. This will pave the way for our ongoing efforts to engineer and transplant more efficient Rubisco into crops.Read moreRead less
Role of a novel zinc-binding motif in the structure-function of deubiquitinating enzymes. The ubiquitin pathway destroys many proteins that control cell function and growth, by attaching ubiquitin to them and marking them for degradation. Deubiquitinating enzymes (DUBs) regulate protein destruction by controlling the amount of ubiquitin attached. DUBs and the ubiquitin pathway can also be manipulated in biotechnology applications. However, very little is known about the structure/function of DUB ....Role of a novel zinc-binding motif in the structure-function of deubiquitinating enzymes. The ubiquitin pathway destroys many proteins that control cell function and growth, by attaching ubiquitin to them and marking them for degradation. Deubiquitinating enzymes (DUBs) regulate protein destruction by controlling the amount of ubiquitin attached. DUBs and the ubiquitin pathway can also be manipulated in biotechnology applications. However, very little is known about the structure/function of DUBs. We have identified a new zinc-binding motif in DUBs, and we will explore how this contributes to their structure, and interactions with other proteins. This will significantly enhance our knowledge of how DUBs function in both biotechnology and in controlling cell function.Read moreRead less
Can efficient algal variants of the photosynthetic CO2-fixing enzyme, Rubisco, be folded and assembled in functional forms in higher-plant plastids? We have shown that it is possible to alter the photosynthetic phenotype of a plant predictably and profoundly by engineering the plastid genome to replace the plant's CO2-fixing enzyme, Rubisco, with a bacterial homolog. Thus it may be possible to replace the plant enzyme with more efficient algal Rubiscos that would allow plants to grow with less l ....Can efficient algal variants of the photosynthetic CO2-fixing enzyme, Rubisco, be folded and assembled in functional forms in higher-plant plastids? We have shown that it is possible to alter the photosynthetic phenotype of a plant predictably and profoundly by engineering the plastid genome to replace the plant's CO2-fixing enzyme, Rubisco, with a bacterial homolog. Thus it may be possible to replace the plant enzyme with more efficient algal Rubiscos that would allow plants to grow with less light, less water or less fertiliser. Before such desirable changes to the plant phenotype can be realised, some complex issues of modification, folding and assembly of Rubisco subunits need to be resolved. This proposal addresses them.Read moreRead less
Directed evolution of enzymes for bioremediation: structure function studies of bimetalloenzymes. We will evolve enzymes that degrade organophosphate pesticides (OPs) that are used in Australian agriculture. Although these OPs were designed to kill insects they are closely related to chemical warfare agents and are known to be toxic to humans. Bacteria have acquired a number of enzymes that degrade some OPs. One such enzyme has been used in field trials demonstrating its potential to degrade OP ....Directed evolution of enzymes for bioremediation: structure function studies of bimetalloenzymes. We will evolve enzymes that degrade organophosphate pesticides (OPs) that are used in Australian agriculture. Although these OPs were designed to kill insects they are closely related to chemical warfare agents and are known to be toxic to humans. Bacteria have acquired a number of enzymes that degrade some OPs. One such enzyme has been used in field trials demonstrating its potential to degrade OP residues. However, many pesticides are not removed rapidly and OP-degrading enzymes require modification(s) if they are to be useful environmental reagents - this can be achieved with directed evolution. Read moreRead less
Active bicarbonate transporters from cyanobacteria: physiological properties, genetic regulation, and introduction into plants for crop improvement. An intriguing set of membrane transport proteins that accumulate bicarbonate into marine cyanobacterial cells will be investigated. These proteins support the crucial process of photosynthetic carbon dioxide fixation in marine cyanobacteria (blue-green algae), which are major contributors to global carbon dioxide sequestration and form one of the f ....Active bicarbonate transporters from cyanobacteria: physiological properties, genetic regulation, and introduction into plants for crop improvement. An intriguing set of membrane transport proteins that accumulate bicarbonate into marine cyanobacterial cells will be investigated. These proteins support the crucial process of photosynthetic carbon dioxide fixation in marine cyanobacteria (blue-green algae), which are major contributors to global carbon dioxide sequestration and form one of the foundations of the marine food web. These bicarbonate "transporters" will also be transferred into a model plant system to test whether the efficiency of photosynthesis can be improved, with corresponding gains in the water-use efficiency of these plants. If successful this technology will have profound global implications for improving crop production in semi-arid areas.Read moreRead less
Directed evolution used to probe protein structure and function; new enzymes for bio-remediation and industry. The aim of the research is to generate new and useful enzymes for bio-remediation and other practical applications. For example, we are evolving enzymes to better degrade organophosphate pesticides that are environmental pollutants. Apart from producing useful enzymes, the proposed research aims at gaining a better understanding of how enzymes work and how they evolve. We intend to dete ....Directed evolution used to probe protein structure and function; new enzymes for bio-remediation and industry. The aim of the research is to generate new and useful enzymes for bio-remediation and other practical applications. For example, we are evolving enzymes to better degrade organophosphate pesticides that are environmental pollutants. Apart from producing useful enzymes, the proposed research aims at gaining a better understanding of how enzymes work and how they evolve. We intend to determine the structure of many related enzymes that have been evolved to have enhanced activities. This data will be used to analyze the intricate relationship between sequence, structure and enzyme activity.Read moreRead less