Purification and characterization of alpha 1-proteinase inhibitor from microsomal fraction of normal human liver.
A plasma alpha 1-proteinase inhibitor was purified, for the first time, from the microsomal fraction of normal human liver by combined procedures of deoxycholate extraction, (NH4)2-SO4 precipitation, gel filtration and immunoadsorbent column chromatographies. The final yield of the material was approx. 15 mg/300 g liver. The preparation was shown to be homogeneous according to polyacrylamide gel electrophoretic and immunological criteria. The estimated molecular weight of the glycoprotein as determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis was 53 000, a value comparable to that reported for plasma alpha 1-proteinase inhibitor. The liver material was, however, devoid of trypsin-, chymotrypsin- or elastase-inhibitory activity. The loss of activity could be due to deoxycholate treatment during the extraction procedure. SDS-polyacrylamide gel electrophoresis of reduced (beta-mercaptoethanol) and alkylated (iodoacetamide) preparation revealed no change in mobility suggesting the presence of only polypeptide chain. The amino acid and carbohydrate compositions of the microsomal material were in close agreement with values reported in the literatures for the corresponding glycoprotein purified from the plasma. The liver microsomal alpha 1-proteinase inhibitor has a single amino terminal residue, namely, glutamic acid, as determined by the dansyl procedure, and a single carboxyl terminal residue, lysine as shown by the carboxypeptidase digestion method. These terminal amino acids are the same as those found in alpha 1-proteinase inhibitor purified from normal human plasma.
D000596:Amino Acids / Q000032:analysis; D002241:Carbohydrates / Q000032:analysis; D002918:Chymotrypsin / Q000037:antagonists & inhibitors; D003902:Detergents / Q000494:pharmacology; D006801:Humans; D008862:Microsomes, Liver / Q000032:analysis*; D008970:Molecular Weight; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors; D000515:alpha 1-Antitrypsin / Q000097:blood / Q000302:isolation & purification* / Q000494:pharmacology
D000596:Amino Acids; D002241:Carbohydrates; D003902:Detergents; D000515:alpha 1-Antitrypsin; D002918:Chymotrypsin; D010196:Pancreatic Elastase