Tesis doctoral de Esther Gratacós Batlle
Carnitine palmitoyltransferase 1 (cpt1) catalyzes the conversion of long chain fatty acyl-coas into acylcarnitines, the first step in the transport of long chain fatty acids from the cytoplasm to the mitochondrial matrix, where they undergo íY-oxidation. This reaction is not only central to the control of fatty acid oxidation, but it also determines the availability of long chain acyl-coa for other processes. There are three different cpt1 isozymes: cpt1a (expressed in liver, pancreas, kidney, brain, blood, and embryonic tissues), cpt1b (expressed only in brown adipose tissue, muscle, and heart) and the recently described cpt1c. cpt1c protein sequence is highly similar to that of the other two isozymes. Expression studies indicate that cpt1c is localized exclusively in the central nervous system, with homogeneous distribution in all areas (hippocampus, cortex, hypothalamus, and others). It has also been reported that cpt1c is localized in neurons but not in astrocytes of adult brain. 1. Cpt1c strucutral model a 3-d strucutral model of the isozyme has been constructed by homology modeling. The residus contacting both substrates have been determined and compared to the same amino acid positions in cpt1a. The results obtained from the analysis show that the residues involved in the catalysis of the reaction in cpt1a and residues contacting both substrates are conserved mainly conserved in cpt1c or show semi-conservative substitutions. 2. Cpt1 enzymatic activity expression of rat cpt1c in saccharomyces cerevisiae yields no catalytic activity when testing different conditions (longer periods of time, increased temperature, increased substrate concentration, testing of microsomal fraction or chimeric protein cpt1·aca). Thus, the yeast expression system is not suitable for studying cpt1c enzymatic activity. 3. Subcellular localization endogenous and overexpressed cpt1c is basically localized in the endoplasmic reticulum of mammalian cells (hek293t, pc12, sh-sy5y, primary cultures of fibroblasts and neurons). Some evidences indicated that cpt1c could also be found, in lower amounts, in mitochondrial associated membranes (mams). the specific sequence of cpt1c n-terminal domain (first 150 amino acids) drives the protein to the endoplasmic reticulum. 4. Cpt1c n-terminus processing the n-terminal end of endogenous cpt1c in wild type mouse brain is processed (at least until val27) and is not detected in mouse brain cortex lysates. 5. Cpt1c membrane topology the n- and c-terminal domains of cpt1c are facing the cytosolic side of the endoplasmic reticulum membrane, whereas the loop domain is facing the endoplasmic reticulum lumen. 6. Cpt1c interacting partners the data provided by the yeast two-hybrid assay do not indicate a unique binding partner of cpt1c. Instead the assay retrieved proteins involved in different functions: protein degradation, membrane trafficking, cell structure, signal transduction and metabolism.
Datos académicos de la tesis doctoral «Molecular characterization of carnitine palmitoyltransferase 1c«
- Título de la tesis: Molecular characterization of carnitine palmitoyltransferase 1c
- Autor: Esther Gratacós Batlle
- Universidad: Barcelona
- Fecha de lectura de la tesis: 16/12/2010
Dirección y tribunal
- Director de la tesis
- Núria Casals Farré
- Tribunal
- Presidente del tribunal: francesc Villarroya gombau
- Miguel Antonio López pérez (vocal)
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