title: Changes in interactions in complexes of hirudin derivatives and human alpha-thrombin due to different crystal forms.
abstract: The three-dimensional structures of D-Phe-Pro-Arg-chloromethyl ketone-inhibited thrombin in complex with Tyr-63-sulfated hirudin (ternary complex) and of thrombin in complex with the bifunctional inhibitor D-Phe-Pro-Arg-Pro-(Gly)4-hirudin (CGP 50,856, binary complex) have been determined by X-ray crystallography in crystal forms different from those described by Skrzypczak-Jankun et al. (Skrzypczak-Jankun, E., Carperos, V.E., Ravichandran, K.G., & Tulinsky, A., 1991, J. Mol. Biol. 221, 1379-1393). In both complexes, the interactions of the C-terminal hirudin segments of the inhibitors binding to the fibrinogen-binding exosite of thrombin are clearly established, including residues 60-64, which are disordered in the earlier crystal form. The interactions of the sulfate group of Tyr-63 in the ternary complex structure explain why natural sulfated hirudin binds with a 10-fold lower K(i) than the desulfated recombinant material. In this new crystal form, the autolysis loop of thrombin (residues 146-150), which is disordered in the earlier crystal form, is ordered due to crystal contacts. Interactions between the C-terminal fragment of hirudin and thrombin are not influenced by crystal contacts in this new crystal form, in contrast to the earlier form. In the bifunctional inhibitor-thrombin complex, the peptide bond between Arg-Pro (P1-P1') seems to be cleaved.
authors.affiliation: Department of Biotechnology, Ciba-Geigy Ltd., Basel, Switzerland.
authors.forename: J P
authors.initials: JP
authors.lastname: Priestle
authors.forename: J
authors.initials: J
authors.lastname: Rahuel
authors.forename: H
authors.initials: H
authors.lastname: Rink
authors.forename: M
authors.initials: M
authors.lastname: Tones
authors.forename: M G
authors.initials: MG
authors.lastname: Grütter
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D004563:Electrochemistry; D005340:Fibrinogen / Q000378:metabolism; D006629:Hirudins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D006860:Hydrogen Bonding; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D013431:Sulfates / Q000378:metabolism; D013917:Thrombin / Q000737:chemistry / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism
chemical_list: D006629:Hirudins; D013431:Sulfates; D014443:Tyrosine; D005340:Fibrinogen; D013917:Thrombin