Journal article
Epitope Randomization Redefines the Functional Role of Glutamic Acid 110 in Interleukin-5 Receptor Activation
The Journal of biological chemistry, v 275(10), pp 7351-7358
10 Mar 2000
PMID: 10702307
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
Sequence randomization through functional phage display of single chain human interleukin (IL)-5 was used to investigate the limits of replaceability of the Glu110 residues that form a part of the receptor-binding epitope. Mutational analysis revealed unexpected affinity for IL-5 receptor α chain with variants containing E110W or E110Y. Escherichia coli-expressed Glu110variants containing E110W in the otherwise sequence-intact N-terminal half, including a variant with an E110A replacement in the sequence-disabled C-terminal half, were shown by their CD spectra to be folded into secondary structures similar to that of single chain human IL-5 (scIL-5). Biosensor kinetics analysis revealed that (E110W/A5)scIL-5 and (E110W/A6)scIL-5 had receptor α chain binding affinities similar to that of (wt/A5)scIL-5. However, (E110W/A6)scIL-5 had a significantly reduced bioactivity in TF-1 cell proliferation compared with both (wt/A5)scIL-5 and (E110W/A5)scIL-5, and this activity reduction was disproportionately greater than the much smaller effect of Glu110 mutation on receptor binding affinity. The marked and disproportionate decrease in TF-1proliferation observed with (E110W/A6)scIL-5 suggests a role for Glu110 in the biological activity mediated by the signal transducing receptor βc subunit of the IL-5 receptor. This is also consistent with the lack of stimulation of JAK2phosphorylation by the (E110W/A6)scIL-5 mutant in recombinant 293T cells, as compared with the concentration-dependent stimulation seen for scIL-5. The results reveal the dispensability of charge in the Glu110 locus of IL-5 for receptor α chain binding and, in contrast, its heretofore underappreciated importance for receptor activation.
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Details
- Title
- Epitope Randomization Redefines the Functional Role of Glutamic Acid 110 in Interleukin-5 Receptor Activation
- Creators
- Sheng-Jiun Wu - University of PennsylvaniaRabindra Tambyraja - University of PennsylvaniaWentao Zhang - University of PennsylvaniaStefan Zahn - University of PennsylvaniaA.Paul Godillot - University of PennsylvaniaIrwin Chaiken - University of Pennsylvania
- Publication Details
- The Journal of biological chemistry, v 275(10), pp 7351-7358
- Publisher
- Elsevier
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Biochemistry and Molecular Biology; Drexel University
- Web of Science ID
- WOS:000085809600087
- Scopus ID
- 2-s2.0-0034629123
- Other Identifier
- 991019520424204721
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- Web of Science research areas
- Biochemistry & Molecular Biology