Probing the interaction of chemokines with their receptors, using NMR spectroscopy
Probing the interaction of chemokines with their receptors, using NMR spectroscopy
Chemokines are a part of the inflammatory response and are involved predominantly in the migration of leukocytes. They have been implicated in diseases such as asthma, dermatitis, rheumatoid arthritis, AIDS and multiple sclerosis. Their receptors are seven transmembrane helix G-coupled proteins which are expressed on the surface of leukocytes. To date over 50 chemokines and 20 receptors have been identified. The disparity between the number of chemokines and receptors is balanced by promiscuity in the ligand-receptor interactions, with most receptors recognising more than one chemokine and several chemokines binding to more than one receptor.
The family of chemokines known as the eotaxin family has three members, CCL11, CCL24 and CCL26 which all activate the receptor CCR3 only. It is hoped that the characterisation of this family’s binding will give a greater insight into the complex chemokine/receptor relationship.
Recombinant CCL24 was expressed in E. coli and labelled with the NMR active isotope 15N before being isolated in a highly purified form.
Peptides to mimic the extracellular loops of the receptor were designed and synthesised. The peptides for extracellular loops 1 and 3 were dissolved in both DMSO and the membrane mimic DPC and their structures solved by NMR spectroscopy. Loop 2 was also investigated, but failed to produce assignable NMR spectra due to reasons unknown.
15N-labelled CCL24 was titrated into loops 1 and 3 suspended in DPC micelles and the resulting systems were studied by NMR spectroscopy. The results gained enabled the building of possible models of the interaction site.
University of Southampton
Long, Joseph Edward
11e2ff49-49ec-445e-a3ba-4405b8a170fd
2004
Long, Joseph Edward
11e2ff49-49ec-445e-a3ba-4405b8a170fd
Long, Joseph Edward
(2004)
Probing the interaction of chemokines with their receptors, using NMR spectroscopy.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
Chemokines are a part of the inflammatory response and are involved predominantly in the migration of leukocytes. They have been implicated in diseases such as asthma, dermatitis, rheumatoid arthritis, AIDS and multiple sclerosis. Their receptors are seven transmembrane helix G-coupled proteins which are expressed on the surface of leukocytes. To date over 50 chemokines and 20 receptors have been identified. The disparity between the number of chemokines and receptors is balanced by promiscuity in the ligand-receptor interactions, with most receptors recognising more than one chemokine and several chemokines binding to more than one receptor.
The family of chemokines known as the eotaxin family has three members, CCL11, CCL24 and CCL26 which all activate the receptor CCR3 only. It is hoped that the characterisation of this family’s binding will give a greater insight into the complex chemokine/receptor relationship.
Recombinant CCL24 was expressed in E. coli and labelled with the NMR active isotope 15N before being isolated in a highly purified form.
Peptides to mimic the extracellular loops of the receptor were designed and synthesised. The peptides for extracellular loops 1 and 3 were dissolved in both DMSO and the membrane mimic DPC and their structures solved by NMR spectroscopy. Loop 2 was also investigated, but failed to produce assignable NMR spectra due to reasons unknown.
15N-labelled CCL24 was titrated into loops 1 and 3 suspended in DPC micelles and the resulting systems were studied by NMR spectroscopy. The results gained enabled the building of possible models of the interaction site.
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Published date: 2004
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Local EPrints ID: 465494
URI: http://eprints.soton.ac.uk/id/eprint/465494
PURE UUID: 4f23ed50-f519-4fb7-af99-8255d8ff7fd4
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Date deposited: 05 Jul 2022 01:26
Last modified: 16 Mar 2024 20:13
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Author:
Joseph Edward Long
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