Cloning and molecular characterization of a human intracellular serine proteinase inhibitor.
We describe a cDNA encoding a serine proteinase inhibitor present in placental tissue and the cytosolic fraction of K562 cells. On the basis of its interaction with thrombin, through which it was discovered, the inhibitor has been operationally named the placental thrombin inhibitor (PTI). Amino acid sequence comparisons suggest that its reactive center is located at Arg-341 and Cys-342, that it lacks a classical N-terminal signal sequence, and that it has the highest degree of similarity to intracellular serine proteinase inhibitors (serpins), such as the human monocyte/neutrophil elastase inhibitor and the equine leukocyte elastase inhibitor. PTI also resembles these inhibitors in that it contains oxidation-sensitive residues adjacent to the reactive site. The PTI cDNA was expressed in rabbit reticulocyte lysate and in COS-7 cells and a 42-kDa protein was produced. Recombinant PTI formed a 67-kDa complex when incubated with thrombin. The ability of native PTI to bind thrombin was destroyed by incubation with iodoacetamide. Analysis of human tissue mRNA indicated that PTI is expressed widely with the highest levels in cardiac and skeletal muscle and placenta. We conclude that PTI is a member of an emerging class of intracellular serpins.
Coughlin
P
P
Department of Medicine, Monash Medical School, Clive Ward Centre, Box Hill Hospital, Australia.
Sun
J
J
Cerruti
L
L
Salem
H H
HH
Bird
P
P
D000595:Amino Acid Sequence; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D007460:Iodoacetamide / Q000737:chemistry; D008969:Molecular Sequence Data; D011994:Recombinant Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015843:Serpins / Q000737:chemistry / Q000235:genetics*; D013917:Thrombin / Q000037:antagonists & inhibitors; D014018:Tissue Distribution
D017931:DNA Primers; D018076:DNA, Complementary; D011994:Recombinant Proteins; D015843:Serpins; D013917:Thrombin; D007460:Iodoacetamide