Isolation, characterization and synthesis of alpha-foetoprotein from neonatal-rat skin.
Monospecific anti-[rat alpha-foetoprotein(alpha-FP)] immunoglobulin G was coupled to CNBr-activated Sepharose-4B (4.5 mg/ml packed volume of gel) to yield an adsorbent. The immunoaffinity column was used to isolate alpha-FP from neonatal-rat skin. Purified skin alpha-FP was found to be immunologically and electrophoretically similar to serum alpha-FP. It yielded a single band with mol.wt. 68000 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. However, on polyacrylamide-gel electrophoresis under non-denaturing conditions, the alpha-FP displayed slow- and fast-moving variants similar to those observed in serum alpha-FP. A Scatchard plot of oestradiol binding to the alpha-FP yielded an association constant of 2.5 X 10(9)M-1 by dextran-coated-charcoal and 0.75 X 10(8)M-1 by Sephadex-gel-filtration procedures respectively. Skin explants from newborn rats were found to incorporate [14C]leucine into immunoprecipitable intracellular alpha-FP. Cycloheximide inhibited the synthesis of alpha-FP in skin explant culture. Our results indicate that newborn-rat skin contains alpha-FP that is similar to serum alpha-FP and which may arise in neonatal-rat skin as a result of synthesis in situ.
D000818:Animals; D000831:Animals, Newborn / Q000378:metabolism*; D002478:Cells, Cultured; D003600:Cytosol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004958:Estradiol / Q000378:metabolism; D005260:Female; D005779:Immunodiffusion; D007122:Immunoelectrophoresis; D007930:Leucine / Q000378:metabolism; D008297:Male; D051381:Rats; D012867:Skin / Q000032:analysis*; D000509:alpha-Fetoproteins / Q000096:biosynthesis / Q000302:isolation & purification* / Q000378:metabolism
D000509:alpha-Fetoproteins; D004958:Estradiol; D007930:Leucine