Protein / Protein Interactions Important For AMP-activated Protein Kinase Regulation
Funder
National Health and Medical Research Council
Funding Amount
$242,545.00
Summary
The AMP-activated protein kinase (AMPK) is an enzyme that monitors the energy levels of the body. When oxygen and nutrient levels decrease, the energy levels of a cell also decrease leading to activation of the AMPK. This results in activation of energy-producing pathways and inhibition of energy-consuming pathways, allowing cells to match supply with demand to ensure their survival. The AMPK comprises of three proteins that together form a functional enzyme. In this application I aim to obtain ....The AMP-activated protein kinase (AMPK) is an enzyme that monitors the energy levels of the body. When oxygen and nutrient levels decrease, the energy levels of a cell also decrease leading to activation of the AMPK. This results in activation of energy-producing pathways and inhibition of energy-consuming pathways, allowing cells to match supply with demand to ensure their survival. The AMPK comprises of three proteins that together form a functional enzyme. In this application I aim to obtain a thorough understanding of the molecular basis of how the AMPK functions. I will determine how and where the three proteins interact with each other and determine where in a cell at any given time the AMPK can be found. This is an important question to answer because many proteins are inactive within the cytoplasm but when they are bound to the plasma membrane they are active. I have previously found the AMPK to be localized to the cytoplasm, membrane and nuclear compartments of the cell, but little is known about the AMPK s function in these different locations. Activation of the AMPK is known to depend on another protein that is also activated when cellular energy levels decrease. This protein has remained elusive to many researchers over the past few years. I plan to identify this protein using new bioinformatics together with the vast amount of information provided by the sequencing of the human genome. Exercise and reduced caloric intake activate the AMPK, these are associated with health benefits and reduce the risk of cardiovascular and neurodegenerative diseases, diabetes and obesity. For these reasons information on the role of the AMPK may improve our understanding of the reasons these diseases develop.Read moreRead less
The Interactions Of Dengue Virus RNA Dependent RNA Polymerase (NS5) With Other Viral And Host Factors.
Funder
National Health and Medical Research Council
Funding Amount
$170,165.00
Summary
Dengue fever is a mosquito-borne disease that is prevalent in tropical countries. It is estimated that 40% of the global population is at risk of dengue infection. Classical dengue fever is not life threatening. However, the more serious disease, dengue haemorrhagic fever-shock syndrome requires intensive medical attention to prevent fatality. A significant number of deaths are recorded each year especially in the underdeveloped countries. Dengue is periodically also a problem in northern Austra ....Dengue fever is a mosquito-borne disease that is prevalent in tropical countries. It is estimated that 40% of the global population is at risk of dengue infection. Classical dengue fever is not life threatening. However, the more serious disease, dengue haemorrhagic fever-shock syndrome requires intensive medical attention to prevent fatality. A significant number of deaths are recorded each year especially in the underdeveloped countries. Dengue is periodically also a problem in northern Australia. There is no cure for dengue fever. The present research aims to use a knowledge-based approach to develop novel antiviral strategies based on preventing the critical protein interactions required for the normal virus life cycle. Two of the most important proteins involved in dengue virus replication are called the NS3 and NS5 proteins. The protein-protein interaction (contact) that occurs between NS5 and NS3 is crucial for the replication of the virus. Little is known about this interaction at present, and the studies we propose will directly address this issue. We have previously shown that a 37 amino acid in the middle of NS5 contains a nuclear localisation signal that can target the normally cytoplasmic protein to the nucleus of the infected cell. What the function of this protein is in the nucleus is not known. We will use a technique called the yeast two-hybrid test to address the question of dengue virus protein interactions in the common bakers yeast. This method is very sensitive and powerful and will provide important insights that will contribute to the development of a rapid high-throughput test to screen the extensive extract collection from Australia's marine biodiversity, held by the Australian Institute of Marine Sciences, to discover suitable inhibitors of NS3-NS5 interaction.Read moreRead less
Studying The Function And Mechanism Of Histone ADP-ribosylation; (ii) Lipo-peptide--protein Vaccine Development
Funder
National Health and Medical Research Council
Funding Amount
$496,076.00
Summary
(i) Histones, which act as DNA supports, are subject to modifications, which may be associated with disease (e.g. cancers). A better understanding of these modifications will provide access to new medicines (ii) The production of new vaccines requires new vaccine development tools. We propose to use fats to produce vaccines which can be delivered via the mouth or nose rather than by injection.
Engineered Histones As DNA Carriers With Application In Therapeutic Gene Delivery
Funder
National Health and Medical Research Council
Funding Amount
$417,750.00
Summary
We intend to apply our knowledge of protein transport to the nucleus to enhance the delivery of DNA to target cells. This relates to the use of gene therapy to treat genetic defects such as inborn errors of metabolism, where a disease-causing lack-of-function mutation can be overcome by engineering cells within the organism which express, in the necessary quantities and in response to the appropriate regulatory signals, the particular component which is lacking. A limiting factor in gene therapy ....We intend to apply our knowledge of protein transport to the nucleus to enhance the delivery of DNA to target cells. This relates to the use of gene therapy to treat genetic defects such as inborn errors of metabolism, where a disease-causing lack-of-function mutation can be overcome by engineering cells within the organism which express, in the necessary quantities and in response to the appropriate regulatory signals, the particular component which is lacking. A limiting factor in gene therapy approaches is the low efficiency of nuclear uptake of introduced DNA, where it has been estimated that < 1% of the DNA taken up is actually expressed. Our proposal seeks to develop approaches to enhance non-viral-mediated gene delivery, in particular by optimising this critical, limiting step of the delivery of exogenous DNA to the nucleus. We intend to apply knowledge from studies of nuclear targeting and chromatin assembly to improve gene transfer technologies. We will build on our work showing that specific signals for nuclear import - nuclear targeting signals (NTSs) - can be used to enhance nuclear gene delivery and expression. Since DNA in the normal cellular context is in the form of chromatin - a specific complex with proteins such as histones - we intend to use reconstituted chromatin as the transfecting DNA, whereby histones engineered to include NTSs and other modular sequence elements will be used. Chromatin should not only enable NTSs and other sequence modules to be linked to the DNA but also protect against nuclease-mediated degradation prior to nuclear entry, condense the DNA to enable more efficient cellular-nuclear entry, and ensure expression of the transfected reporter gene by presenting it to the cell in a physiological context. Our approaches should contribute to bringing gene therapy closer to reality in the clinic.Read moreRead less