Characterization of a cDNA coding for human protein C.

Abstract

Protein C is a precursor to a serine protease that is present in mammalian plasma. In its activated form, it readily inactivates factor Va and factor VIIIa, 2 proteins that participate as cofactors in the blood coagulation cascade. A .lambda.gt11 library containing cDNA [complementary DNA] inserts prepared from human liver mRNA was screened with an antibody to human protein C. Seven positive clones were isolated from 2 .times. 106 phage and were plaque-purified. The cDNA inserts of 2 of these phage were sequenced and shown to code for human protein C. Each cDNA insert coded for a portion of the light chain of the molecule, a connecting region, the heavy chain, a stop codon, a 3''-noncoding region and a poly(A) tail. The length of the noncoding sequence on the 3'' end differed in the 2 clones, but each contained a processing or polyadenylylation signal followed by a poly(A) tail. The amino acid sequence as determined from the cDNA indicates that protein C is synthesized as a single-chain polypeptide containing the light chain and the heavy chain connected by a dipeptide of Lys-Arg. The single-chain molecule is then converted to the light and heavy chains by cleavage of 2 or more internal peptide bonds. In plasma, the heavy and light chains of protein C are linked together by a disulfide bond. The amino acid seqeunce of human protein C shows a high degree of homology with that of the bovine molecule. The DNA sequence coding for the catalytic region near the active site serine in human protein C also showed a high degree of DNA and amino acid sequence identity with prothrombin, factor IX and factor X, 3 of the other vitamin K-dependent serine proteases that are present in plasma.