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Kinetic analysis of the interleukin-13 receptor complex

Kinetic analysis of the interleukin-13 receptor complex
Kinetic analysis of the interleukin-13 receptor complex
Interleukin (IL)-13 is a key cytokine associated with the asthmatic phenotype. It signals via its cognate receptor, a complex of IL-13 receptor ?1 chain (IL-13R?1) with IL-4R?; however, a second protein, IL-13R?2, also binds IL-13. To determine the binding contributions of the individual components of the IL-13 receptor to IL-13, we have employed surface plasmon resonance and equilibrium binding assays to investigate the ligand binding characteristics of shIL-13R?1, shIL-13R?2, and IL-4R?. shIL-13R?1 bound IL-13 with moderate affinity (KD = 37.8 ± 1.8 nM, n = 10), whereas no binding was observed for hIL-4R?. In contrast, shIL-13R?2 produced a high affinity interaction with IL-13 (KD = 2.49 ± 0.94 nM n = 10). IL-13R?2 exhibited the binding characteristics of a negative regulator with a fast association rate and an exceptional slow dissociation rate. Although IL-13 interacted weakly with IL-4R? on its own (KD > 50 µM), the presence of hIL-4R? significantly increased the affinity of shIL-13R?1 for IL-13 but had no effect on the binding affinity of IL-13R?2. Detailed kinetic analyses of the binding properties of the heteromeric complexes suggested a sequential mechanism for the binding of IL-13 to its signaling receptor, in which IL-13 first binds to IL-13R?1 and this then recruits IL-4? to stabilize a high affinity interaction.
Abbreviations: IL, interleukin; shIL-13R?1, soluble human IL-13 receptor ? chain 1; shIL-13R?2, soluble human IL-13 receptor ? chain 2; sIL-4R?, soluble human IL-4 receptor; SPR, surface plasmon resonance, RU resonance units; STAT, signal transducers and activators of transcription; SA chip, streptavidin-coated sensor chip.
0021-9258
46073-46078
Andrews, Allison-Lynn
4ddaec43-0f43-40cd-a191-aa53b0b30f16
Holloway, John W.
4bbd77e6-c095-445d-a36b-a50a72f6fe1a
Puddicombe, Sarah M.
124e2c4e-ab9a-46f3-855c-b54ed0b61cc4
Holgate, Stephen T.
2e7c17a9-6796-436e-8772-1fe6d2ac5edc
Davies, Donna E.
7de8fdc7-3640-4e3a-aa91-d0e03f990c38
Andrews, Allison-Lynn
4ddaec43-0f43-40cd-a191-aa53b0b30f16
Holloway, John W.
4bbd77e6-c095-445d-a36b-a50a72f6fe1a
Puddicombe, Sarah M.
124e2c4e-ab9a-46f3-855c-b54ed0b61cc4
Holgate, Stephen T.
2e7c17a9-6796-436e-8772-1fe6d2ac5edc
Davies, Donna E.
7de8fdc7-3640-4e3a-aa91-d0e03f990c38

Andrews, Allison-Lynn, Holloway, John W., Puddicombe, Sarah M., Holgate, Stephen T. and Davies, Donna E. (2002) Kinetic analysis of the interleukin-13 receptor complex. The Journal of Biological Chemistry, 277 (48), 46073-46078. (doi:10.1074/jbc.M209560200).

Record type: Article

Abstract

Interleukin (IL)-13 is a key cytokine associated with the asthmatic phenotype. It signals via its cognate receptor, a complex of IL-13 receptor ?1 chain (IL-13R?1) with IL-4R?; however, a second protein, IL-13R?2, also binds IL-13. To determine the binding contributions of the individual components of the IL-13 receptor to IL-13, we have employed surface plasmon resonance and equilibrium binding assays to investigate the ligand binding characteristics of shIL-13R?1, shIL-13R?2, and IL-4R?. shIL-13R?1 bound IL-13 with moderate affinity (KD = 37.8 ± 1.8 nM, n = 10), whereas no binding was observed for hIL-4R?. In contrast, shIL-13R?2 produced a high affinity interaction with IL-13 (KD = 2.49 ± 0.94 nM n = 10). IL-13R?2 exhibited the binding characteristics of a negative regulator with a fast association rate and an exceptional slow dissociation rate. Although IL-13 interacted weakly with IL-4R? on its own (KD > 50 µM), the presence of hIL-4R? significantly increased the affinity of shIL-13R?1 for IL-13 but had no effect on the binding affinity of IL-13R?2. Detailed kinetic analyses of the binding properties of the heteromeric complexes suggested a sequential mechanism for the binding of IL-13 to its signaling receptor, in which IL-13 first binds to IL-13R?1 and this then recruits IL-4? to stabilize a high affinity interaction.
Abbreviations: IL, interleukin; shIL-13R?1, soluble human IL-13 receptor ? chain 1; shIL-13R?2, soluble human IL-13 receptor ? chain 2; sIL-4R?, soluble human IL-4 receptor; SPR, surface plasmon resonance, RU resonance units; STAT, signal transducers and activators of transcription; SA chip, streptavidin-coated sensor chip.

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Published date: 2002

Identifiers

Local EPrints ID: 26907
URI: http://eprints.soton.ac.uk/id/eprint/26907
ISSN: 0021-9258
PURE UUID: f9c5caae-e784-40c0-8a1b-b54437fc4fd0
ORCID for John W. Holloway: ORCID iD orcid.org/0000-0001-9998-0464
ORCID for Donna E. Davies: ORCID iD orcid.org/0000-0002-5117-2991

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Date deposited: 25 Apr 2006
Last modified: 18 Feb 2021 16:49

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