Design and characterization of hirulogs: A novel class of bivalent peptide inhibitors of thrombin

Journal Article · Tue Jul 31 00:00:00 EDT 1990 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00482a021· OSTI ID:6249554

Maraganore, J M; Bourdon, P; Jablonski, J; Ramachandran, K L ^[1]; Fenton, II, J W ^[2]

---

1. Biogen, Inc., Cambridge, MA (USA)
2. New York State Department of Health, Albany (USA)

A novel class of synthetic peptides has been designed that inhibit the thrombin catalytic site and exhibit specificity for the anion-binding exosite (ABE) of {alpha}-thrombin. These peptides, called hirulogs, consist of (i) an active-site specificity sequence with a restricted Arg-Pro scissile bond, (ii) a polymeric linker of glycyl residues from 6 to 18 {angstrom} in length, and (iii) an ABE recognition sequence such as that in the hirudin C-terminus. Hirulog-1 ((D-Phe)-Pro-Arg-Pro-(Gly){sub 4}-Asn-Gly-Asp-Phe-Glu-Glu-Ile-Pro-Glu-Tyr-Leu) inhibits the thrombin-catalyzed hydrolysis of a tripeptide p-nitroanilide substrate with K{sub i} = 2.3 nM. In contrast, the synthetic C-terminal hirudin peptide S-Hir{sub 53-64}, which binds to the thrombin ABE, blocked the fibrinogen clotting activity of the enzyme with K{sub i} = 144 nM but failed to inhibit the hydrolysis of p-nitroanilide substrates at concentrations as high as 1 mM. Hirulog-1, but not S-Hir{sub 53-64}, was found to inhibit the incorporation of ({sup 14}C)diisopropyl fluorophosphate in thrombin. Hirulog-1 appears specific for thrombin as it lacks inhibitory activities toward human factor Xa, human plasmin, and bovine trypsin at inhibitor:enzyme concentrations 3 orders of magnitude higher than those required to inhibit thrombin. The optimal inhibitory activity of hirulog-1 depends upon all three components of its structure. Comparison of anticoagulant activities of hirulog-1, hirudin, and S-Hir{sub 53-64} showed that the synthetic hirulog-1 is 2-fold more potent than hirudin and 100-fold more active than S-Hir{sub 53-64} in increasing the activated partial thromboplastin time of normal human plasma.

• OSTI ID: 6249554
• Journal Information: Biochemistry; (USA), Vol. 29:30; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
ANTICOAGULANTS
MOLECULAR STRUCTURE
CARBON 14 COMPOUNDS
UPTAKE
PEPTIDES
BIOCHEMICAL REACTION KINETICS
THROMBIN
INHIBITION
CATTLE
GLYCINE
THROMBOSIS
TRYPSIN
AMINO ACIDS
ANIMALS
CARBOXYLIC ACIDS
CARDIOVASCULAR DISEASES
COAGULANTS
DISEASES
DOMESTIC ANIMALS
DRUGS
ENZYMES
HEMATOLOGIC AGENTS
HEMOSTATICS
HYDROLASES
KINETICS
LABELLED COMPOUNDS
MAMMALS
ORGANIC ACIDS
ORGANIC COMPOUNDS
PEPTIDE HYDROLASES
PROTEINS
REACTION KINETICS
RUMINANTS
SERINE PROTEINASES
VASCULAR DISEASES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques