Mammalian ADP-dependent glucokinase : a thesis presented in partial fulfilment of the requirement for the degree of Master of Science in Biochemistry at Massey University, Palmerston North, New Zealand
The mammalian ADP-dependent glucokinase is the most recent mammalian glucokinase to have been discovered, and is unique in its ability to catalyse the phosphorylation of glucose to glucose-6-phosphate using ADP as the phosphoryl donor. Up until the discovery of this enzyme, the traditional biochemical view was that the first step of glycolysis was solely catalysed by ATP-dependent hexokinases, types I-IV. The particular role played by ADP-GK in the mammalian cell and the significance of this role has not yet been determined, although it is hypothesised that the ADP-dependent glucokinase could be potentially significant in contributing to the survival of cells under low energy and hypoxic or ischemic conditions. By using ADP as the energy investment in phase one of the glycolytic cycle instead of ATP, it is predicted that glycolysis could be sustained for longer during lower energy conditions (conditions of high ADP:ATP ratios). Since the phosphorylation of glucose by ADP-GK results in the production of AMP, it may also be possible that this has a direct effect on the energy charge of the cell. The AMP produced could lead to the regulation of cellular metabolism during hypoxia and/or ischemia via the activation of the cell-energy regulator AMPK. The study of mammalian ADP-dependent glucokinase is a very new area, and prior to this no investigation of the human ADP-GK enzyme had been undertaken. The main objective of this project was to clone, express and purify the recombinant ADP-GK so it could be kinetically characterised and directly compared with the recombinant mouse kinetic characteristics, the only other mammalian ADP-GK to have been studied. Unfortunately, due to complications in the expression and purification of soluble recombinant human ADP-GK, the project did not incorporate the kinetic characterisation of the enzyme. Acquiring data on the kinetic characteristics of the human ADP-GK will, in the long term, assist in the elucidation of the metabolic role of this enzyme, so the continuation of this project would be worthwhile.