Molecular basis for the inhibition of human alpha-thrombin by the macrocyclic peptide cyclotheonamide A.
The macrocyclic peptide cyclotheonamide A (CtA), isolated from the marine sponge Theonella sp., represents an unusual class of serine protease inhibitor. A complex of this inhibitor with human alpha-thrombin, a protease central to the bioregulation of thrombosis and hemostasis, was studied by x-ray crystallography. This work (2.3-A resolution) confirms the structure of CtA and reveals intimate details about its molecular recognition within the enzyme active site. Interactions due to the "Pro-Arg motif" (Arg occupancy of the S1 specificity pocket; formation of a hydrogen-bonded two-strand antiparallel beta-sheet with Ser214-Gly216) and the alpha-keto amide group of CtA are primarily responsible for binding to thrombin, with the alpha-keto amide serving as a transition-state analogue. A special interaction with the "insertion loop" of thrombin (Tyr60A-Thr60I) is manifested through engagement of the hydroxyphenyl group of CtA with Trp60D as part of an "aromatic stacking chain." Biochemical inhibition data ( Ki values at 37 degrees C) were obtained for CtA with thrombin and a diverse collection of serine proteases. Thus, CtA is just a moderate inhibitor of human alpha-thrombin ( Ki = 0.18 microM) but a potent inhibitor of trypsin ( Ki = 0.023 microM) and streptokinase ( Ki = 0.035 microM). The relative lack of potency of CtA as a thrombin inhibitor is discussed with respect to certain structural features of the enzyme complex. We also report the total synthesis of CtA, by a convergent [2 + 3] fragment-condensation approach, to serve the preparation of cyclotheonamide analogues for structure-function studies.[1]References
- Molecular basis for the inhibition of human alpha-thrombin by the macrocyclic peptide cyclotheonamide A. Maryanoff, B.E., Qiu, X., Padmanabhan, K.P., Tulinsky, A., Almond, H.R., Andrade-Gordon, P., Greco, M.N., Kauffman, J.A., Nicolaou, K.C., Liu, A. Proc. Natl. Acad. Sci. U.S.A. (1993) [Pubmed]
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