The cDNAs encoding two isoforms, S (slow) and F (fast), of alpha 1-antiproteinase (also referred to as alpha 1-antitrypsin or alpha 1-proteinase inhibitor) as well as contrapsin were obtained by screening lambda gt11 cDNA library prepared fro inflamed guinea pig liver. The sequence analyses of these cDNAs and NH2-terminal peptides of the purified proteins revealed that both isoforms of alpha 1-antiproteinase consist of 405 amino acid residues including a signal peptide of 24 residues and that contrapsin consists of 410 amino acid residues with the same length of the signal peptide. Guinea pig contrapsin had 89, 88, 62, 42, and 41% homology to its own alpha 1-antiproteinases F and S, rat alpha 1-antiproteinase, mouse and rat contrapsins, respectively. This suggests that guinea pig contrapsin is not orthologous to mouse and rat contrapsins and that it developed from a much later duplication of alpha 1-antiproteinase gene after the guinea pig had diverged from the murine lineage. The available data suggest that the reactive site region of alpha 1-antiproteinase can be categorized into orthodox and unorthodox types: the former has P3-P'3 consensus sequence of Xaa-Pro-Met-Ser-Xaa-Pro, where Xaa is Leu, Ile, Val, or Met, while the latter, which occurs in species having multiple alpha 1-antiproteinase isoforms, has the sequence whose P1 Met has changed to other amino acids. Thus, the reactive site region of the orthodox type, which occurs in all seven mammals examined to date, is highly conserved. This is in marked contrast to the fact that the same region is hypervariable among the paralogous proteins belonging to the serpin superfamily.