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Isotopic labelling and heteronuclear multidimensional NMR methods for the structural study of the protein complex, Lactobacillus casei dihydrofolate reductase with methotrexate

Soteriou, Alice; (1995) Isotopic labelling and heteronuclear multidimensional NMR methods for the structural study of the protein complex, Lactobacillus casei dihydrofolate reductase with methotrexate. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Dihydrofolate reductase (DHFR) is an enzyme which is a target for a number of anticancer, antibacterial and antimalarial drugs. There are many unanswered questions concerning the interactions between DHFR and its ligands, for example why some specifically bind to bacterial DHFRs rather than mammalian species. In order to attempt to answer these questions it is necessary to obtain structural information about such complexes in solution. High resolution heteronuclear multidimensional Nuclear Magnetic Resonance (NMR) spectroscopy has been used to study the structure of the complex of Lactobacillus casei DHFR with the anticancer drug, methotrexate. Isotopic labelling (13C and 15N) and of the protein has been combined with the appropriate multidimensional method to provide the necessary spectral simplification to make assignments in the protein-ligand complex. Firstly the resonances according to residue type have been determined from -TOCSY and -COSY based techniques and secondly. NOESY-based experiments have provided the distance information which allows particular residues to be located within the protein molecules. As well as finding the global molecular folds of the protein, specific torsion angle information has been reported. The latter data is important in assessing the local geometry of each residue. Using the data from the above, structure calculations have been carried out using simulated annealing (molecular dynamics) techniques to generate a three- dimensional structure of the protein-ligand complex, in solution.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Isotopic labelling and heteronuclear multidimensional NMR methods for the structural study of the protein complex, Lactobacillus casei dihydrofolate reductase with methotrexate
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
Keywords: Health and environmental sciences; Heteronuclear multidimensional NMR; Isotopic labelling
URI: https://discovery.ucl.ac.uk/id/eprint/10102537
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