Murine soluble Fc gamma receptors/IgG-binding factors (IgG-BF): analysis of the relation to Fc gamma RII and production of milligram quantities of biologically active recombinant IgG-BF.
The production of soluble forms of low-affinity Fc gamma R by cells expressing recombinant or natural membrane Fc gamma RII, and the structural relationships between these soluble receptors and membrane Fc gamma RII are described. We show that 37-40 kD soluble Fc gamma RII, corresponding to the two N-terminal domains of Fc gamma RII and binding to IgG, are spontaneously produced in vitro by cleavage of membrane Fc gamma RII. Moreover, we describe methods to produce and purify to homogeneity large quantities of endotoxin-free recombinant IgG-binding factor (rIgG-BF) from the culture medium of a cell line transfected with a mutated Fc gamma RII cDNA. These methods include the use of bioreactors for culturing transfected fibroblasts and the purification of rIgG-BF by ion-exchange chromatography and hydrophobic-interaction chromatography. By using such procedures, about 2.4 mg of rIgG-BF were purified from 1 liter of culture medium of transfected fibroblasts. Like natural IgG-BF, the 95-99% pure rIgG-BF suppressed, in a dose-dependent manner, secondary in vitro IgG antibody responses to sheep red blood cells.
Sautès
C
C
INSERM U. 255, Institut Curie, Paris, France.
Mazières
N
N
Galinha
A
A
Tartour
E
E
Bonnerot
C
C
Amigorena
S
S
Teillaud
C
C
Spagnoli
R
R
Fridman
W H
WH
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006031:Glycosylation; D007074:Immunoglobulin G / Q000096:biosynthesis; D008222:Lymphokines / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D029584:Prostatic Secretory Proteins; D017452:Receptors, IgG / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012995:Solubility; D013491:Suppressor Factors, Immunologic / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D014162:Transfection
D007074:Immunoglobulin G; D008222:Lymphokines; D029584:Prostatic Secretory Proteins; D017452:Receptors, IgG; D011994:Recombinant Proteins; D013491:Suppressor Factors, Immunologic; C043076:beta-microseminoprotein; C013059:immunoglobulin-binding factors