Interleukin-1 receptor antagonist in inflammatory exudate cells of rabbits. Production, purification and determination of primary structure.
A rabbit interleukin-1 (IL-1) inhibitor in inflammatory peritoneal exudate cells was purified to apparent homogeneity. This inhibitor was extracted from exudate cells of the 24-hr stage of casein-induced peritoneal inflammation and purified using isoelectrofocusing (IEF), gel filtration, followed in this order by high-performance liquid chromatography (HPLC) steps with hydroxylapatite and anionic ion exchanger. The purified factor showed a single band on silver-stained SDS-PAGE. This molecule of MW 19,000 and pI 5.5 inhibited the binding of both IL-1 alpha and beta to receptors on a thymoma cell line, EL-4 and a B-cell line, 70Z/3. We determined its primary structure by a combination of peptide chemistry and molecular cloning. The inhibitor was synthesized as a precursor composed of 177 amino acids and was processed to a mature molecule of 143 amino acids. The N-terminal amino acid of the mature inhibitor was N-acetyl-methionine residue. The deduced amino acid sequence of the inhibitor showed a 77% homology to the human IL-1 receptor antagonist (IL-1Ra) and essentially the same mode of action as seen with human IL-1Ra. We consider that this inhibitor is a rabbit counterpart of human IL-1Ra, although there are differences with respect to the molecular structure; the N-terminus of the mature rabbit IL-1Ra at a position of nine amino acids downstream from that of human IL-1Ra.
Goto
F
F
Department of Pathology, Kumamoto University Medical School, Japan.
Goto
K
K
Miyata
T
T
Ohkawara
S
S
Takao
T
T
Mori
S
S
Furukawa
S
S
Maeda
T
T
Iwanaga
S
S
Shimonishi
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001202:Ascitic Fluid / Q000276:immunology*; D001483:Base Sequence; D001685:Biological Factors / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D007525:Isoelectric Focusing; D008297:Male; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011817:Rabbits / Q000276:immunology*; D017472:Receptors, Interleukin-1 / Q000037:antagonists & inhibitors*
D001685:Biological Factors; D010446:Peptide Fragments; D017472:Receptors, Interleukin-1