Biomaterial Database

The isomorphous structures of prethrombin2, hirugen-, and PPACK-thrombin: changes accompanying activation and exosite binding to thrombin. 1994

J Vijayalakshmi, and K P Padmanabhan, and K G Mann, and A Tulinsky

Department of Chemistry, Michigan State University, East Lansing 48824-1322, USA.

The X-ray crystal structure of prethrombin2 (pre2), the immediate inactive precursor of alpha-thrombin, has been determined at 2.0 A resolution complexed with hirugen. The structure has been refined to a final R-value of 0.169 using 14,211 observed reflections in the resolution range 8.0-2.0 A. A total of 202 water molecules have also been located in the structure. Comparison with the hirugen-thrombin complex showed that, apart from the flexible beginning and terminal regions of the molecule, there are 4 polypeptide segments in pre2 differing in conformation from the active enzyme (Pro 186-Asp 194, Gly 216-Gly 223, Gly 142-Pro 152, and the Arg 15-Ile 16 cleavage region). The formation of the Ile 16-Asp 194 ion pair and the specificity pocket are characteristic of serine protease activation with the conformation of the catalytic triad being conserved. With the determination of isomorphous structures of hirugen-thrombin and D-Phe-Pro-Arg chloromethyl ketone (PPACK)-thrombin, the changes that occur in the active site that affect the kinetics of chromogenic substrate hydrolysis on binding to the fibrinogen recognition exosite have been determined. The backbone of the Ala 190-Gly 197 segment in the active site has an average RMS difference of 0.55 A between the 2 structures (about 3.7 sigma compared to the bulk structure). This segment has 2 type II beta-bends, the first bend showing the largest shift due to hirugen binding. Another important feature was the 2 different conformations of the side chain of Glu 192. The side chain extends to solvent in hirugen-thrombin, which is compatible with the binding of substrates having an acidic residue in the P3 position (protein-C, thrombin platelet receptor). In PPACK-thrombin, the side chain of Asp 189 and the segment Arg 221A-Gly 223 move to provide space for the inhibitor, whereas in hirugen-thrombin, the Ala 190-Gly 197 movement expands the active site region. Although 8 water molecules are expelled from the active site with PPACK binding, the inhibitor complex is resolvated with 5 other water molecules.

UI	MeSH Term	Description	Entries
D008958	Models, Molecular	Models used experimentally or theoretically to study molecular shape, electronic properties, or interactions; includes analogous molecules, computer-generated graphics, and mechanical structures.	Molecular Models,Model, Molecular,Molecular Model
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D010446	Peptide Fragments	Partial proteins formed by partial hydrolysis of complete proteins or generated through PROTEIN ENGINEERING techniques.	Peptide Fragment,Fragment, Peptide,Fragments, Peptide
D011485	Protein Binding	The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments.	Plasma Protein Binding Capacity,Binding, Protein
D011487	Protein Conformation	The characteristic 3-dimensional shape of a protein, including the secondary, supersecondary (motifs), tertiary (domains) and quaternary structure of the peptide chain. PROTEIN STRUCTURE, QUATERNARY describes the conformation assumed by multimeric proteins (aggregates of more than one polypeptide chain).	Conformation, Protein,Conformations, Protein,Protein Conformations
D011516	Prothrombin	A plasma protein that is the inactive precursor of thrombin. It is converted to thrombin by a prothrombin activator complex consisting of factor Xa, factor V, phospholipid, and calcium ions. Deficiency of prothrombin leads to hypoprothrombinemia.	Coagulation Factor II,Factor II,Blood Coagulation Factor II,Differentiation Reversal Factor,Factor II, Coagulation,Factor, Differentiation Reversal,II, Coagulation Factor
D004789	Enzyme Activation	Conversion of an inactive form of an enzyme to one possessing metabolic activity. It includes 1, activation by ions (activators); 2, activation by cofactors (coenzymes); and 3, conversion of an enzyme precursor (proenzyme or zymogen) to an active enzyme.	Activation, Enzyme,Activations, Enzyme,Enzyme Activations
D004792	Enzyme Precursors	Physiologically inactive substances that can be converted to active enzymes.	Enzyme Precursor,Proenzyme,Proenzymes,Zymogen,Zymogens,Precursor, Enzyme,Precursors, Enzyme
D006629	Hirudins	Single-chain polypeptides of about 65 amino acids (7 kDa) from LEECHES that have a neutral hydrophobic N terminus, an acidic hydrophilic C terminus, and a compact, hydrophobic core region. Recombinant hirudins lack tyr-63 sulfation and are referred to as 'desulfato-hirudins'. They form a stable non-covalent complex with ALPHA-THROMBIN, thereby abolishing its ability to cleave FIBRINOGEN.	Hirudin
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man