Purification of homogeneity and amino acid sequence analysis of a receptor protein for interleukin 1.
The interleukin 1 (IL-1) receptor from mouse EL-4 thymoma cells was purified to homogeneity by a method which utilized ligand affinity chromatography and classical chromatographic techniques. After solubilization of the receptor from intact cells with the zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, the IL-1 binding activity was purified greater than 23,000-fold. Analysis of the purified protein by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblot, and ligand blot demonstrated that a single protein of molecular mass of approximately 80 kDa is the IL-1 binding polypeptide. The purified protein bound IL-1 with a dissociation constant of approximately 1.1 X 10(-10) M, which is indistinguishable from the affinity of the cell-bound receptor. The amino acid composition of this protein is strikingly similar to the composition deduced from the sequence of a cDNA coding for an IL-1 receptor from EL-4 cells. Protein sequence analysis of Staphylococcus aureus V-8 protease-derived peptides yields data consistent with the sequence proposed from cloned cDNA. These studies have demonstrated that the high affinity IL-1 receptor on EL-4 cells is the 80-kDa protein.
Stern
A S
AS
Department of Protein Biochemistry, Roche Research Center, Hoffmann-LaRoche, Inc., Nutley, New Jersey 07110.
Pan
Y C
YC
Hellmann
R S
RS
Parker
K P
KP
Mueller
D
D
Hulmes
J D
JD
Kilian
P L
PL
Chizzonite
R
R
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002460:Cell Line; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D007375:Interleukin-1 / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D008970:Molecular Weight; D011948:Receptors, Antigen, T-Cell / Q000302:isolation & purification*; D011971:Receptors, Immunologic / Q000302:isolation & purification* / Q000378:metabolism; D017472:Receptors, Interleukin-1; D013945:Thymoma / Q000276:immunology; D013953:Thymus Neoplasms / Q000276:immunology
D000596:Amino Acids; D007375:Interleukin-1; D011948:Receptors, Antigen, T-Cell; D011971:Receptors, Immunologic; D017472:Receptors, Interleukin-1