Recombinant soluble CD59 inhibits reactive haemolysis with complement.
Three soluble forms of membrane attack complex inhibitory factor (MACIF or CD59) were prepared using recombinant baculovirus-infected insect cells. They consisted of 70, 77 and 86 amino acids, starting from the amino terminus of naturally occurring CD59, and were designated recombinant (r) CD59 70, 77 and 86, respectively. All three rCD59 lacked a glycosyl-phosphatidylinositol (GPI) anchor, unlike membrane CD59 which has a GPI anchor at the anchor at the carboxyl terminus (77th amino acid). Their activities in inhibiting complement activation were assayed with C5b-7 intermediate cells and C8 and C9 components. The inhibitory activity of rCD59 70 was as high as that of rCD59 77 and twice that of rCD59 86. In addition, it was one-fourth and one-hundredth lower than the activities of urine and erythrocyte CD59, respectively. However, when assayed in the presence of human serum at a final concentration of 50% (v/v), the activities of both urine and erythrocyte CD59 were greatly decreased to to one-tenth of that of rCD59 70. Purified rCD59 70 molecules were all glycosylated, but rCD59 77 and 86 were mixtures of glycosylated and non-glycosylated molecules. The inhibitory activities of rCD59 77 and 86 were the same for the glycosylated and non-glycosylated forms. These results suggest that the soluble rCD59 provide a means for elucidating the biological roles of CD59.
Sugita
Y
Y
Molecular Medicine Research Laboratories, Yamanouchi Pharmaceutical Co. Ltd, Tsukuba, Japan.
Ito
K
K
Shiozuka
K
K
Suzuki
H
H
Gushima
H
H
Tomita
M
M
Masuho
Y
Y
D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000737:chemistry / Q000276:immunology* / Q000652:urine; D001483:Base Sequence; D018957:CD59 Antigens; D003165:Complement System Proteins / Q000276:immunology*; D004306:Dose-Response Relationship, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000276:immunology; D006461:Hemolysis / Q000276:immunology*; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000276:immunology* / Q000652:urine; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology; D012995:Solubility; D013329:Structure-Activity Relationship
D015703:Antigens, CD; D018957:CD59 Antigens; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D003165:Complement System Proteins