Isolation and characterization of hirustasin, an antistasin-type serine-proteinase inhibitor from the medical leech Hirudo medicinalis.
Antistasin, a potent inhibitor of the blood coagulation factor Xa, is the prototype of a novel family of serine-proteinase inhibitors. We have now isolated, sequenced and characterized an antistasin-type inhibitor from the medical leech Hirudo medicinalis. Hirustasin (Hirudo antistasin) was purified to apparent homogeneity by cation-exchange and affinity chromatography. Amino acid sequencing of the 55 amino acid protein (M(r) 5866) revealed that hirustasin is the only antistasin-type protein known to consist of one domain only; 27% and 32% sequence identity was found to the first and second domains of antistasin, respectively, and a nearly exact conservation of the spacing of the ten cysteine residues. Hirustasin is the first inhibitor of tissue kallikrein identified in leeches, and is also a tight-binding inhibitor of trypsin, chymotrypsin and neutrophil cathepsin G. However, despite the high similarity to antistasin, particularly in the vicinity of the putative reactive-site peptide bond, hirustasin neither inhibits blood coagulation in vitro nor amidolytic activity of isolated factor Xa. Thus, structural elements other than the reactive site sequence significantly influence the specificity of antistasin-type proteinase inhibitors.
Söllner
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Abteilung für Klinische Chemie und Klinische Biochemie, Klinikum Innenstadt, Ludwig-Maximilians-Universität, München, Germany.
Mentele
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Eckerskorn
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Fritz
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Sommerhoff
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CP
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D005591:Chemical Fractionation; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D007447:Invertebrate Hormones / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D007865:Leeches / Q000737:chemistry*; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D015842:Serine Proteinase Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D013045:Species Specificity
D000596:Amino Acids; D007447:Invertebrate Hormones; D015842:Serine Proteinase Inhibitors; C053155:antistasin; C086021:hirustasin