Phage presentation and affinity selection of a deletion mutant of human lnterleukin-3 |
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Authors: | Sean Merlin Edwin Rowold Ann Abegg Cathleen Berglund Jon Klover Nick Staten John P McKearn Stephen C Lee |
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Institution: | (1) Searle Research and Development, Monsanto Company, 700 Chesterfield Parkway, 63198 St. Louis, MO, USA |
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Abstract: | A deletion derivative of the cytokine human interleukin-3 (hIL-315–125, comprising amino acids 15–125 of the native protein) was produced as a fusion to the filamentous phage surface protein pIII.
The cytokine was detected in association with phage particles by protein immunoblotting. Compared to an equivalent quantity
of soluble cytokine, phage-presented hIL-315–125 exhibited reduced biological activity in a hIL-3-dependent cell proliferation assay. The reduction in activity was attributable
to presence of phage particles in the assay, rather than directly owing to physical incorporation of the cytokine into the
phage particle. Owing to the position of the amber codon in the phagemid vector, the phagemid-produced free hIL-315–125 species (designated hIL-315–125 ε) had 20 amino acids appended to its C-terminus; hIL-315–125 ε did not exhibit reduced bioactivity. hIL-315–125-presenting phage were affinity-selected with either a hIL-3-reactive polyclonal antibody or with cells expressing the heterodimeric
hIL-3 receptor. These data are consistent with the use of phage-display technology for the affinity selection of hIL-3 variants
with modified biological properties. |
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Keywords: | Phage display cytokine receptor growth factor proliferation biopanning |
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