Liver mitochondrial aldehyde dehydrogenase is coded by a nuclear gene and after being synthesized on a ribosome must be transported across the two mitochondrial membranes. It is known that these matrix space mitochondrial enzymes are initially synthesized as a higher molecular weight precursor with an N-terminal amino acid extension. After being imported into the mitochondria the extension, called a signal peptide or leader sequence, is removed by a specific protease. Little is known about the actual three dimensional structure of a signal peptide.
The aim of this proposal is use 2-dimensional nuclear magnetic resonance to solve the structures of synthetic peptides corresponding to the signal peptide from human, beef and rat lever aldehyde dehydrogenase. This will be done in solution as well as when the peptide is bound to a micelle in order to simulate what could be expected to be found when the peptide is bound to a membrane. Specific amino acids in the rat enzyme's signal peptide will be changed by performing site directed mutagenesis. Peptides corresponding to the signal peptide of those mutants which are not imported will be made and their structure determined. The goal is to understand the role of structure in the import process. It has recently been shown that ethanol inhibits the translocation, hence structures will be determined in its presence as well as in the presence of helix-stabilizing compounds. Transport of proteins into organelles is a vital cell function. Knowing the structure of the signal peptide may allow for the development of a drug to inhibit the process. In addition, if alcohol is found to alter the conformation of the peptide it will indicate that an additional consequence of abusive drinking is to decrease the cell's ability to transport proteins.
| Thornton, K; Wang, Y; Weiner, H et al. (1993) Import, processing, and two-dimensional NMR structure of a linker-deleted signal peptide of rat liver mitochondrial aldehyde dehydrogenase. J Biol Chem 268:19906-14 |
| Wang, Y; Weiner, H (1993) The presequence of rat liver aldehyde dehydrogenase requires the presence of an alpha-helix at its N-terminal region which is stabilized by the helix at its C termini. J Biol Chem 268:4759-65 |
| Karslake, C; Piotto, M E; Pak, Y K et al. (1990) 2D NMR and structural model for a mitochondrial signal peptide bound to a micelle. Biochemistry 29:9872-8 |
