TY - JOUR
T1 - Requirement of hydrophilic amino-terminal residues for granulocyte-macrophage colony-stimulating factor bioactivity and receptor binding
AU - Meropol, Neal J.
AU - Altmann, Scott W.
AU - Shanafelt, Armen B.
AU - Kastelein, Robert A.
AU - Johnson, G. Douglas
AU - Prystowsky, Michael B.
PY - 1992/7/15
Y1 - 1992/7/15
N2 - Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a glycoprotein required for the proliferation and differentiation of granulocyte and macrophage precursors. Previous investigations have identified regions in human and murine GM-CSF that are required for bioactivity. In the present study, alanine substitution mutagenesis was undertaken to define more precisely specific amino-terminal residues in murine GM-CSF that are involved in bioactivity and receptor binding. Five double alanine mutants were identified that showed at least 10-fold reductions in bioactivity (K14AK20A, K14AE21A, H15AK20A, H15AE21A, K20AE21A). Each of these mutants maintained a normal N-linked glycosylation pattern when expressed in COS-1 cells, suggesting that native polypeptide backbone conformation was preserved. The purified prokaryotic expression products of two mutants (K14AE21A and H15AE21A) had a 100-fold decrease in bioactivity and a decrease in receptor binding, indicating that the side chains of K14, H15, and E21 are required for optimal receptor binding and maximal bioactivity.
AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a glycoprotein required for the proliferation and differentiation of granulocyte and macrophage precursors. Previous investigations have identified regions in human and murine GM-CSF that are required for bioactivity. In the present study, alanine substitution mutagenesis was undertaken to define more precisely specific amino-terminal residues in murine GM-CSF that are involved in bioactivity and receptor binding. Five double alanine mutants were identified that showed at least 10-fold reductions in bioactivity (K14AK20A, K14AE21A, H15AK20A, H15AE21A, K20AE21A). Each of these mutants maintained a normal N-linked glycosylation pattern when expressed in COS-1 cells, suggesting that native polypeptide backbone conformation was preserved. The purified prokaryotic expression products of two mutants (K14AE21A and H15AE21A) had a 100-fold decrease in bioactivity and a decrease in receptor binding, indicating that the side chains of K14, H15, and E21 are required for optimal receptor binding and maximal bioactivity.
UR - http://www.scopus.com/inward/record.url?scp=0026716429&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0026716429&partnerID=8YFLogxK
M3 - Article
C2 - 1385812
AN - SCOPUS:0026716429
SN - 0021-9258
VL - 267
SP - 14266
EP - 14269
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 20
ER -