Regulation of pyruvate dehydrogenase kinase 4 (PDK4) through protein-protein interaction and its role in apoptosis
Date
2009-10-20T08:02:30Z
Authors
Ntlabati, Patricia Coliwe
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Abstract
Pyruvate dehydrogenase kinase 4 (PDK4), a mammalian mitochondrial serine kinase has emerged as an interesting candidate for diabetes therapy. Due to the high prevalence of this disease especially type 2 diabetes (T2D) and the health complications associated with it, there is extensive effort to find the appropriate treatment. Understanding the regulation of PDK4 would therefore contribute significantly to the development of therapeutic agents.
This research outlines the bioinformatics analysis of PDK4, using tools such as Interweaver, ClustalW and Protein Structure Visualiser. These programs were used to determine potentially interacting partners for PDK4. Interweaver database identified 96 proteins which have possible interaction sites for PDK4. Protein p100/p49, containing a death domain that is known to have a role in suppressing apoptosis, was identified as a potential partner for PDK4. The alignment between p100/p49 primary sequence and that of PDK4 using Gene explorer/ClustalW demonstrated sequence similarity between the two proteins. Swiss PDB Viewer then located the positions of the amino acids that are in the hypothetical protein binding motif of p100/p49 within the 3D structure of hPDK4. These amino acids were found to be located in the region of PDK4 which is known to bind protein substrates of PDK4 and may be accessible to other proteins as well. These findings were very interesting as PDK4 have not previously been associated with apoptosis and this could be the link between apoptosis and insulin resistance. Cell biology studies were then performed to verify the relationship between PDK4 and apoptosis. In this regard, HeLa and HepG2 cells were treated with apoptosis inducing agents such as TNFα, C2-Ceramide, and Linoleic acid. These cells were then monitored for apoptosis and PDK4 mRNA expression using a DNA laddering assay as well as Real Time PCR. The results showed that these factors to induce apoptosis in a concentration dependent manner. This induction of apoptosis by these three factors indicated to suppress PDK4 mRNA levels. These findings suggested a regulatory role for PDK4 in apoptosis.