Purification and partial characterization of a preneoplastic antigen in liver carcinogenesis.
A preneoplastic antigen has been isolated from the microsomal fraction of the hyperplastic nodules of rat liver. During the purification, the presence of preneoplastic antigen was monitored by microcomplement fixation test and double immunodiffusion plate following reaction with antiserum specific for this antigen. The antigen was purified by seven steps with a purification factor of 167 and 11% yield. It migrated as a single protein band, both in nondenaturing and denaturing polyacrylamide gel electrophoresis; showed a single sharp peak in an isoelectric focusing column at pH 4.75; and had a single serine amino-terminal amino acid. In the absence of detergent during purification, this protein tends to form molecular aggregates. It appears to be a glycoprotein based on the positive periodic acid-Schiff stain and absorption by concanavalin A:Sepharose affinity column chromatography. Amino acid analysis indicated that the protein is acidic, having acidic amino acids (aspartate and glutamate) and basic amino acids (lysine, arginine, and histidine) in a ratio of 6.8. An estimate of the molecular weight by gel filtration gave a value of 145,000 and a calculated Stokes radius of 41.5 A. Electrophoresis under denaturing conditions on sodium dodecyl sulfate-polyacrylamide gel indicated that this protein is a dimer consisting of two subunits of equal molecular weight (M.W. 74,000). Circular dichroism spectra indicated alpha-helix contents at 298 and 222 nm of 25.5 and 28.3%, respectively. A similar protein is present in the endoplasmic reticulum of normal rat liver, but in a much lower concentration.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000951:Antigens, Neoplasm / Q000032:analysis / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D007525:Isoelectric Focusing; D008113:Liver Neoplasms / Q000276:immunology*; D008297:Male; D011230:Precancerous Conditions; D011863:Radioimmunoassay; D051381:Rats
D000596:Amino Acids; D000951:Antigens, Neoplasm