Identification of residues critical to the activity of human granulocyte colony-stimulating factor

John F. Reidhaar-Olson, Janet A. De Souza-Hart, Harold E. Selick

Research output: Contribution to journalArticle

42 Citations (Scopus)

Abstract

Alanine scanning mutagenesis of human granulocyte colony-stimulating factor (G-CSF) was used to identify residues critical for the cell- proliferative activity of the protein. Fifty-eight residues, most of them on the protein surface, were independently mutated to alanine. Most of the variants retained full biological activity; however, 15 mutants were significantly impaired in their ability to stimulate bone marrow cell proliferation in vitro. Four of these variants contain mutations at buried residues and two have substitutions at side chains involved in intramolecular hydrogen bonds. The remaining nine down mutations identify two regions on the surface of the molecule important for biological activity. Consistent with these observations, measurements of binding to NFS-60 cells indicate that the residues most important for receptor binding are Lys40 and Phe144 in site 1 and Glu19 in site 2. In addition to these residues, Val48 and Leu49 in site 1 and Leu15, Asp112, and Leu124 in site 2 are also important for biological activity. These results suggest the presence of two binding sites on the cytokine surface required for dimerization of the G-CSF receptor.

Original languageEnglish (US)
Pages (from-to)9034-9041
Number of pages8
JournalBiochemistry
Volume35
Issue number28
DOIs
StatePublished - 1996
Externally publishedYes

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Granulocyte Colony-Stimulating Factor
Bioactivity
Human Activities
Alanine
Granulocyte Colony-Stimulating Factor Receptors
Mutation
Dimerization
Mutagenesis
Bone Marrow Cells
Hydrogen
Membrane Proteins
Binding Sites
Cell Proliferation
Cell proliferation
Cytokines
Hydrogen bonds
Bone
Substitution reactions
Scanning
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

Identification of residues critical to the activity of human granulocyte colony-stimulating factor. / Reidhaar-Olson, John F.; De Souza-Hart, Janet A.; Selick, Harold E.

In: Biochemistry, Vol. 35, No. 28, 1996, p. 9034-9041.

Research output: Contribution to journalArticle

Reidhaar-Olson, John F. ; De Souza-Hart, Janet A. ; Selick, Harold E. / Identification of residues critical to the activity of human granulocyte colony-stimulating factor. In: Biochemistry. 1996 ; Vol. 35, No. 28. pp. 9034-9041.
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