Molecular cloning and structural characterization of the hagfish proteinase inhibitor of the alpha-2-macroglobulin family.
The "most primitive" living vertebrate the hagfish has a dimeric proteinase inhibitor, a protein homologous to human alpha2-macroglobulin, in its plasma at high concentration. Although the hagfish proteinase inhibitor has been isolated and its function and quaternary structure studied, its primary structure, subunit composition and fragmentation process remain unclear. In this study, hagfish proteinase inhibitor cDNA was cloned, sequenced and cDNA-deduced amino acid sequence was analyzed. A large fraction of homosubunits in the dimeric structure of the protein has undergone a cleavage at a specific arginyl residue (Arg833) while the rest retained their chain integrity without being processed. Thus random combinations of processed and nonprocessed subunits in the dimeric structure of this protein result in different molecular conformers and generate a complicated multiband pattern in SDS-PAGE. It was further demonstrated by proteolytic analysis that the hagfish inhibitor has no susceptible arginyl residues within its bait region and thus incapable of trapping arginine specific proteinases. This implies that the specific subunit cleavage at Arg833 was caused by an unknown arginine specific proteinase which escaped from the entrapment by the hagfish inhibitor.
Laboratory of Biodynamics, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
Alimjan
A
Idiris
Ken-ichi
K
Ohtsubo
Koh-ichi
K
Yoza
Toshiya
T
Osada
Noboru
N
Nakamichi
Toshiharu
T
Matsumura
Atsushi
A
Ikai
D000510:Alpha-Globulins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006196:Hagfishes / Q000097:blood / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D021122:Protein Subunits; D020539:Sequence Analysis, Protein
D000510:Alpha-Globulins; D018076:DNA, Complementary; D011480:Protease Inhibitors; D021122:Protein Subunits; C025500:alpha(2)-microglobulin