Amino acid metabolism in mammalian brain: discovery and characterisation of mammalian ketimine reductase

There are subtle differences in amino acid metabolism in mammalian brain, especially as regards lysine degradation. In lysine degradation the pipecolate pathway predominates in adult mammalian brain whereas the saccharopine pathway predominates in the rest of the body. This suggests that this pathway may play an important role in brain development and functioning. A key enzyme involved in the pipecolate pathway is a ketimine reductase which is involved in the reduction of Δ¹-piperideine-2-carboxylate to form L-pipecolate. This enzyme is also involved in proline metabolism in the reduction of Δ¹-pyrroline-2-carboxylate to form L-proline. In addition the enzyme is involved in the reduction of cyclic ketimines originating from sulfur-containing amino acid metabolism. Prior to this research no mammalian ketimine reductases had been identified in mammalian genomes. The enzyme was identified as the protein μ-crystallin (CRYM). It was found to be strongly inhibited regulated by thyroid hormones and may regulate enzyme activity in vivo. A novel catalytic mechanism is also suggested based on in silico docking where an active site arginine residue acts as a proton donor. Prior to this research it was not known as an enzyme, however it was known as an important thyroid hormone binding protein and a structural protein in diurnal marsupial lens. This multi-disciplinary research project involved organic synthesis of enzyme substrates, classical biochemical enzyme purification, proteomic identification, in silico docking, bioinformatics, and kinetic analysis of activity and inhibition. This research was self-initiated and this thesis consists of three research papers and two reviews which have been published or submitted during the course of my candidature with myself as primary author. Publications which I have also been involved in as a minor co-author and a letter to the editor are included in the appendices. This research is an important contribution to our understanding of amino acid metabolism in the mammalian brain, and also its relationship with thyroid hormones.