A study of the binding of anti-aggregatory substances to platelet bound plasma proteins.

Simpson, Christopher

Abstract

The binding of a number of platelet active compounds to serum albumin was studied using several direct fluorescence methods and fluorescence polarisation. Particular emphasis was placed on the study of the binding of dipyridamole (DPD) and its analogues. The data obtained from the above experiments was compared with that obtained from the classical methods of equilibrium dialysis, dynamic dialysis, and continuous ultrafiltration and used to set up a number of nonlinear least squares regression analysis computer programmes to analyse binding isotherms. In the course of this study, the fluorescence methods were applied to a number of binding systems not previously investigated. Several binding systems showed no change in the fluorescence characteristics of the protein or ligand. This led to the development of a Sephadex batch method to be used in conjunction with fluorescence. The concentration of free ligand was measured directly in the Sephadex gel. Factors which effect the fluorescence from the Sephadex bed were extensively studied. To show the flexibility of this method it was applied to a number of binding systems involving both high and low affinity constants and to the binding of basic and acidic compounds. Fluorescence methods were, for the first time, applied to the binding of a number of ligands to OC1 acid glycoprotein (AGP). Using classical and fluorescence methods some of the characteristics of the binding site were established. Extensive use was made of competitive studies as part of this work. Having established binding characteristics of a number of platelet active ligands to plasma proteins studies were conducted with platelets. A number of methods for washing platelets were investigated in order to find an efficient and reproducible method of washing platelets. The binding of DPD to platelets was then studied using direct fluorescence, fluorescence polarisation and a centrifuging difference method. Two classes of binding sites were observed and so the significance of both the primary and secondary sites were investigated. Possible correlation of the binding with a number of enzyme and uptake systems was attempted together with a comparison of the binding with that of AGP. It was shown that the primary site correlates with the inhibition of adenosine uptake and the secondary with the inhibition of phosphodiesterase activity. As part of this work the binding of DPD was compared with the binding of a number of other ligands. Finally the importance of the binding in terms of the inhibition of aggregation by DPD was discussed.

Citation

SIMPSON, C. 1983. A study of the binding of anti-aggregatory substances to platelet bound plasma proteins. Robert Gordon's Institute of Technology, PhD thesis. Hosted on OpenAIR [online]. Available from: https://doi.org/10.48526/rgu-wt-1993218

• Thesis Type: Thesis
• Deposit Date: Oct 2, 2024
• Publicly Available Date: Oct 2, 2024
• DOI: https://doi.org/10.48526/rgu-wt-1993218
• Public URL: https://rgu-repository.worktribe.com/output/1993218
• Award Date: Nov 30, 1983