Purification to homogeneity of pig leucocyte catabolin, a protein that causes cartilage resorption in vitro.
Catabolin, a protein that causes proteoglycan resorption in explants of living cartilage, was purified to homogeneity from culture medium conditioned by culturing buffy-coat leucocytes from 60 litres of pig blood in the presence of concanavalin A. The purification steps were (1) gel filtration of concentrated medium, (2) chromatofocusing, (3) hydroxyapatite chromatography, (4) anion-exchange chromatography (Mono Q), (5) reversed-phase high-pressure liquid chromatography (h.p.l.c.) (Zorbax ODS). These achieved approx. 9000-fold purification from the starting material. The purified protein when reduced ran as a single band on sodium dodecyl sulphate (SDS)/polyacrylamide-gel electrophoresis with Mr 21000. On isoelectric focusing its pI was 4.8-5.0, and there was evidence of micro-heterogeneity. The protein co-migrated with active material on h.p.l.c., isoelectric focusing and SDS gels (15 and 12.5% acrylamide) under both reducing and non-reducing conditions. The pure protein caused proteoglycan release from cultured bovine nasal cartilage at 20pM. Its possible identity with interleukin 1 is discussed.
D000818:Animals; D002356:Cartilage / Q000187:drug effects; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D007375:Interleukin-1; D053583:Interleukin-1beta; D007525:Isoelectric Focusing; D007962:Leukocytes / Q000032:analysis*; D011506:Proteins / Q000302:isolation & purification* / Q000494:pharmacology; D013552:Swine
D007375:Interleukin-1; D053583:Interleukin-1beta; D011506:Proteins