Máquinas moleculares que sintetizan anhidridos fosfóricos.
This thesis is about four enzymes that synthesize phosphoric anhydrides, and which have key roles on the biosynthetic pathways of ornithine/arginine, proline, lysine and isoleucine, and on the microbial synthesis of pyrimidine nucleotides. A novel and characteristic fold common to the enzymes carbamate kinase (CK) and acetylglutamate kinase (NAGK) had been identified by other members of the laboratory, previously to my incorporation. CK and NAGK belong to the amino acid kinase structural family (PFAM database, PF00696; http://www.sanger.ac.uk/Software/Pfam), that also includes aspartokinase (AK), glutamate 5-kinase, bacterial UMP kinase and N-acetyl L-glutamate synthase. Based on the sequence similarity of these enzymes, the laboratory proposed a common fold for all of them, equivalent to that previously described for CK and NAGK. One of the main objectives of the laboratory and of my work has been to probe this hypothesis experimentally. Initially, I studied NAGK from E. coli and I corroborated by site-directed mutagenesis, functional inferences derived from the previous structural work. The high similitude between NAGK and AK allowed to perform a site-directed mutagenesis study of AK, and to use the results of this study to build a molecular model of the AKIII from E. coli. The third enzyme that I have studied, UMP kinase, is a key enzyme of the pathway of biosynthesis of pyrimidine nucleotides in bacteria, that different to the other enzymes of the amino acid kinase family, synthetizes a phosphoric-phosphoric anhydride. I have determined the 3D-structure of the UMP kinase from Pyrococcus furiosus using X-ray crystallography, and also of the glutamate 5-kinase fom E. coli, which is a key enzyme of the biosynthesic pathway of proline in microorganisms and plants, and also of ornithine in animals. The work of this thesis is included in the following publications:
- Marco-Marín, C. et al (2003). Journal of Molecular Biology (2003) 334, 459-476.
- Marco-Marín, C. et al (2005). Biochim. Biophys. Acta. (2005) 1747, 271-275.
- Marco-Marín, C. et al (2005). Journal of Molecular Biology (2005) 352, 438-454.
- Marco-Marín, C. et al (2007). Journal of Molecular Biology (2007) 367, 1431-1446.
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Advisor:Rubio Zamora, Vicente
School:Universitat de València
Source Type:Master's Thesis
Keywords:bioquímica i biologia molecular
Date of Publication:02/19/2007