Comparison of nerve growth factor receptor binding models using heterodimeric muteins
| Autor: | Hrishikesh M. Mehta, Kenneth E. Neet, Sang B. Woo |
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| Rok vydání: | 2012 |
| Předmět: |
Models
Molecular animal structures Neurite Cell Survival Plasma protein binding Biology Ligands Heteroreceptor Binding Competitive PC12 Cells Receptor Nerve Growth Factor Article Cell Line Mice Cellular and Molecular Neuroscience Nerve Growth Factor Protein Interaction Mapping Neurites Animals Low-affinity nerve growth factor receptor Phosphorylation Receptor trkA Binding site Receptor Binding Sites Cell Differentiation Fibroblasts Protein-Tyrosine Kinases Nerve growth factor receptor binding Molecular biology Rats Cell biology Nerve growth factor nervous system Mutation Dimerization Protein Processing Post-Translational Protein Binding |
| Zdroj: | Journal of Neuroscience Research. 90:2259-2271 |
| ISSN: | 0360-4012 |
| DOI: | 10.1002/jnr.23116 |
| Popis: | Nerve growth factor (NGF) is a homodimer that binds to two distinct receptor types, TrkA and p75, to support survival and differentiation of neurons. The high-affinity binding on the cell surface is believed to involve a heteroreceptor complex, but its exact nature is unclear. We developed a heterodimer (heteromutein) of two NGF muteins that can bind p75 and TrkA on opposite sides of the heterodimer, but not two TrkA receptors. Previously described muteins are Δ9/13 that is TrkA negative and 7-84-103 that is signal selective through TrkA. The heteromutein (Htm1) was used to study the heteroreceptor complex formation and function, in the putative absence of NGF-induced TrkA dimerization. Cellular binding assays indicated that Htm1 does not bind TrkA as efficiently as wild-type (wt) NGF but has better affinity than either homodimeric mutein. Htm1, 7-84-103, and Δ9/13 were each able to compete for cold-temperature, cold-chase stable binding on PC12 cells, indicating that binding to p75 was required for a portion of this high-affinity binding. Survival, neurite outgrowth, and MAPK signaling in PC12 cells also showed a reduced response for Htm1, compared with wtNGF, but was better than the parent muteins in the order wtNGF > Htm1 > 7-84-103 >> Δ9/13. Htm1 and 7-84-103 demonstrated similar levels of survival on cells expressing only TrkA. In the longstanding debate on the NGF receptor binding mechanism, our data support the ligand passing of NGF from p75 to TrkA involving a transient heteroreceptor complex of p75-NGF-TrkA. |
| Databáze: | OpenAIRE |
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