Physicochemical characterization of lymphocyte-activating factor (LAF).
The LAF produced by the mouse macrophage cell line, P388D1, is a single polypeptide chain of m.w. 12,000 to 16,000 daltons. Native LAF was destroyed by Streptomyces griseus protease, but not by trypsin, chymotrypsin, and papain, although in the presence of 8 M urea, papain completely destroyed LAF activity. LAF did not bind to concanavalin A-Sepharose, suggesting that LAF does not contain significant amounts of mannosyl or glycosyl residues. Since LAF activity was not inactivated by a treatment of reduction and alkylation the active conformation of LAF does not appear to be dependent on disulfide linkages. LAF was not irreversibly denatured by 8 M urea or 0.1 to 0.5% SDS. On SDS-polyacrylamide gels, the m.w. of LAF was 12,000 daltons, as compared to a value of 16,000 daltons, as determined by gel filtration. The isoelectric point of LAF was 5.0 to 5.4 as determined on 7.5% acrylamide gels (pH 3 to 10). On the basis of these results it appears that the P388D1 cell line-derived LAF is a relatively stable molecule that shares several physicochemical properties with normal human and mouse macrophage-derived LAF.
D000478:Alkylation; D000818:Animals; D001665:Binding Sites; D055598:Chemical Phenomena; D002621:Chemistry; D002918:Chymotrypsin / Q000494:pharmacology; D003208:Concanavalin A / Q000494:pharmacology; D007376:Interleukin-2; D008214:Lymphocytes / Q000276:immunology*; D008222:Lymphokines; D051379:Mice; D010206:Papain / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000494:pharmacology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology; D014508:Urea / Q000494:pharmacology
D007376:Interleukin-2; D008222:Lymphokines; D003208:Concanavalin A; D012967:Sodium Dodecyl Sulfate; D014508:Urea; D010447:Peptide Hydrolases; D002918:Chymotrypsin; D014357:Trypsin; D010206:Papain