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Title: Structural studies of immunophilin-ligand complexes
Author: Yuande, Y.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2003
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The work described in this thesis used crystallographic and biochemical methods to study the complex of immunophins (humans Cyclophilin A (hCypA) and FK506 binding protein (FKBP)) with a series of chemically synthesised ligands. HCypA is the cytosolic receptor of the immunosuppressant Cyclosporin A (CsA), and FKBP is the receptor for the immunosuppressant FK506. Both hCypA and FKBP possess peptidyl-prolyl isomerization (PPIase) enzymatic activity. In this project hCypA was produced and characterised in house. SDS page electrophoresis, mass spectrometry and PPIase enzymatic assays were used to characterise hCypA, which was expressed in E. coli and purified in house with chromatographic techniques. PPIase assays and fluorescence spectrometry were used to identify nine novel ligand complexes. The ligands were synthesised in the Chemistry department of Edinburgh University. Crystal complexes of the nine hCypA ligand complexes were obtained and crystal structures were solved up to a resolution of 1.8 A. It was observed that all nine ligands bind hCypA in a similar mode, and the relationship between the binding strength and structural features was evaluated. The following equation was developed to relate measured dissociation constant (Kd) and structural features: Kd = 238 - 0.185 * (Buried area) = 17.9* (Number of Hydrogen bonds less than 3A) - 6.7 * (Number of Hydrogen bonds greater than 3.0 A) FKBP was also crystallised. The complex crystal of FKBP and the dipeptide leucylproline (Leu-Pro) was prepared. The crystal structure of FKBP and Leu-Pro shows that the amide bond of bound Leu-Pro is twisted and provides an insight into the PPIase mechanism.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available