Thrombin inhibition by cyclic peptides from thrombomodulin.
Peptides corresponding to the loop regions of the fourth, fifth, and sixth epidermal growth factor (EGF)-like domains of thrombomodulin (TM) have been synthesized and assayed for thrombin inhibition, as indicated by both inhibition of thrombin-mediated fibrinogen clotting and inhibition of the association of thrombin with TM that results in protein C activation. Peptides from the fifth EGF-like domain showed significant inhibition of fibrinogen clotting and protein C activation, whereas peptides from the fourth and sixth EGF-like domains were weak inhibitors in both assays. Two structural features were important for inhibitory potency of the peptides from the fifth EGF-like domain: cyclization by a disulfide bond and attachment of the "tail" amino acids C-terminal to the disulfide loop. Linear control peptides did not significantly inhibit clotting or protein C activation. The C-terminal loop alone, the "tail" peptide, or a mixture of the two were at least 10-fold less potent inhibitors of clotting or protein C activation. A more constrained peptide analog was designed by deletion of an isoleucine within the C5-C6 disulfide loop, TM52-1 + 5C. This analog was a better inhibitor in both assay systems, having a Ki for protein C activation of 26 microM.
Lougheed
J C
JC
Department of Chemistry and Biochemistry, University of California-San Diego, La Jolla 92093-0601, USA.
Bowman
C L
CL
Meininger
D P
DP
Komives
E A
EA
D000595:Amino Acid Sequence; D001665:Binding Sites; D001777:Blood Coagulation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010456:Peptides, Cyclic / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D011485:Protein Binding; D011486:Protein C / Q000037:antagonists & inhibitors / Q000378:metabolism; D013917:Thrombin / Q000037:antagonists & inhibitors* / Q000378:metabolism; D018180:Thrombomodulin / Q000737:chemistry* / Q000378:metabolism
D010456:Peptides, Cyclic; D011486:Protein C; D018180:Thrombomodulin; D004815:Epidermal Growth Factor; D013917:Thrombin