The areas around the three phosphorylation sites are shown in red. Site 1 is in the bottom left corner, site 2 in the top right, and site 3 in the bottom right.
PDK thus participates in the regulation of thepyruvate dehydrogenase complex of which pyruvate dehydrogenase is the first component. Both PDK and the pyruvate dehydrogenase complex are located in themitochondrial matrix ofeukaryotes. The complex acts to convertpyruvate (a product ofglycolysis in thecytosol) toacetyl-coA, which is then oxidized in the mitochondria to produce energy, in thecitric acid cycle. Bydownregulating the activity of this complex, PDK will decrease the oxidation of pyruvate in mitochondria and increase the conversion of pyruvate tolactate in the cytosol.
PDK can phosphorylate aserine residue onpyruvate dehydrogenase at three possible sites. Some evidence has shown that phosphorylation at site 1 will nearly completely deactivate the enzyme while phosphorylation at sites 2 and 3 had only a small contribution to complex inactivation.[1] Therefore, it is phosphorylation at site 1 that is responsible for pyruvate dehydrogenase deactivation.
The primary sequencing between the four isozymes are conserved with 70% identity. The greatest differences occur near theN-terminus.[2]
PDK1 is the largest of the four with 436residues while PDK2, PDK3 and PDK4 have 407, 406, and 411 residues respectively. The isozymes have different activity and phosphorylation rates at each site. At site 1 in order from fastest to slowest, PDK2 > PDK4 ≈ PDK1 > PDK3. For site 2, PDK3 > PDK4 > PDK2 > PDK1. Only PDK1 can phosphorylate site 3. However, it has been shown that these activities are sensitive to slight changes inpH so the microenvironment of the PDK isozymes may change the reaction rates.[3][4]
Isozyme abundance has also been shown to betissue specific. PDK1 is ample inheart cells. PDK3 is most abundant intestis. PDK2 is present in most tissues but low inspleen andlung cells. PDK4 is predominantly found inskeletal muscle andheart tissues.[5]
Pyruvate dehydrogenase is deactivated when phosphorylated by PDK. Normally, the active site of pyruvate dehydrogenase is in a stabilized and ordered conformation supported by a network ofhydrogen bonds. However, phosphorylation by PDK at site 1 causessteric clashes with another nearby serine residue due to both the increased size and negative charges associated with the phosphorylated residue.[6] This disrupts the hydrogen bond network and disorders the conformation of two phosphorylation loops. These loops prevent the reductiveacetylation step, thus halting overall activity of the enzyme.[7] The conformational changes and mechanism of deactivation for phosphorylation at sites 2 and 3 are not known at this time.
Each isozyme responds to each of these factors slightly differently. NADH stimulates PDK1 activity by 20% and PDK2 activity by 30%. NADH with acetyl-CoA increases activity in these enzymes by 200% and 300% respectively. In similar conditions, PDK3 is unresponsive to NADH and inhibited by NADH with acetyl-CoA. PDK4 has a 200% activity increase with NADH, but adding acetyl-CoA does not increase activity further.[5]
PDK isoforms are elevated in obesity, diabetes, heart failure, and cancer.[10] Some studies have shown that cells that lackinsulin (or are insensitive to insulin) overexpress PDK4.[11] As a result, the pyruvate formed from glycolysis cannot be oxidized which leads tohyperglycaemia due to the fact that glucose in the blood cannot be used efficiently. Therefore, several drugs target PDK4 hoping to treattype II diabetes.[12]
PDK1 has shown to have increased activity inhypoxic cancer cells due to the presence ofHIF-1. PDK1 shunts pyruvate away from the citric acid cycle and keeps the hypoxic cell alive.[13] Therefore, PDK1 inhibition has been suggested as an antitumor therapy since PDK1 preventsapoptosis in these cancerous cells.[14] Similarly, PDK3 has been shown to be overexpressed in colon cancer cell lines.[15] Three proposed inhibitors are AZD7545 anddichloroacetate which both bind to PDK1, andRadicicol which binds to PDK3.[16]
^Yeaman SJ, Hutcheson ET, Roche TE, Pettit FH, Brown JR, Reed LJ, Watson DC, Dixon GH (June 1978). "Sites of phosphorylation on pyruvate dehydrogenase from bovine kidney and heart".Biochemistry.17 (12):2364–70.doi:10.1021/bi00605a017.PMID678513.
^Sugden MC, Holness MJ (May 2003). "Recent advances in mechanisms regulating glucose oxidation at the level of the pyruvate dehydrogenase complex by PDKs".American Journal of Physiology. Endocrinology and Metabolism.284 (5): E855-62.doi:10.1152/ajpendo.00526.2002.PMID12676647.
^Holness MJ, Sugden MC (December 2003). "Regulation of pyruvate dehydrogenase complex activity by reversible phosphorylation".Biochemical Society Transactions.31 (Pt 6):1143–51.doi:10.1042/bst0311143.PMID14641014.
^Meurs KM, Lahmers S, Keene BW, White SN, Oyama MA, Mauceli E, Lindblad-Toh K (August 2012). "A splice site mutation in a gene encoding for PDK4, a mitochondrial protein, is associated with the development of dilated cardiomyopathy in the Doberman pinscher".Human Genetics.131 (8):1319–25.doi:10.1007/s00439-012-1158-2.PMID22447147.S2CID253975177.
