PDBsum entry 1mh0

Blood clotting

PDB id: 1mh0

Contents

Protein chain

277 a.a. *

Ligands

NAG ×2

Waters ×111

* Residue conservation analysis

PDB id:

1mh0

Name:

Blood clotting

Title:

Crystal structure of the anticoagulant slow form of thrombin

Structure:

Prothrombin. Chain: a, b. Synonym: alpha-thrombin. Coagulation factor ii. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Organ: blood. Expressed in: cricetulus griseus. Expression_system_taxid: 10029. Expression_system_organ: kidney. Expression_system_tissue: kidney cells.

Resolution:

2.80Å R-factor: 0.219 R-free: 0.275

Authors:

A.O.Pineda,S.Savvides,G.Waksman,E.Di Cera

Key ref:

A.O.Pineda et al. (2002). Crystal structure of the anticoagulant slow form of thrombin. J Biol Chem, 277, 40177-40180. PubMed id: 12205081 DOI: 10.1074/jbc.C200465200

Date:

18-Aug-02 Release date: 08-Nov-02

Protein chains

P00734  (THRB_HUMAN) -  Prothrombin from Homo sapiens

Seq: 622 a.a.

Struc: 277 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.4.21.5 - thrombin.
• Reaction: Preferential cleavage: Arg-|-Gly; activates fibrinogen to fibrin and releases fibrinopeptide A and B.

DOI no: 10.1074/jbc.C200465200

J Biol Chem 277:40177-40180 (2002)

PubMed id: 12205081

Crystal structure of the anticoagulant slow form of thrombin.

A.O.Pineda, S.N.Savvides, G.Waksman, E.Di Cera.

ABSTRACT

Using the thrombin mutant R77aA devoid of the site of autoproteolytic degradation at exosite I, we have solved for the first time the structure of thrombin free of any inhibitors and effector molecules and stabilized in the Na(+)-free slow form. The slow form shows subtle differences compared with the currently available structures of the Na(+)-bound fast form that carry inhibitors at the active site or exosite I. The most notable differences are the displacement of Asp-189 in the S1 specificity pocket, a downward shift of the 190-193 strand, a rearrangement of the side chain of Glu-192, and a significant shift in the position of the catalytic Ser-195 that is no longer within H-bonding distance from His-57. The structure of the slow form explains the reduced specificity toward synthetic and natural substrates and suggests a molecular basis for its anticoagulant properties.

Selected figure(s)

Figure 1.
Fig. 1. Backbone traces of thrombin in the slow (blue) and fast (yellow) forms. The root mean square deviation between the two structures is 0.5 Å. The structure of the slow form refers to molecule A in the asymmetric unit. The structures are portrayed in the "Bode orientation" (11) with the active site at the center, the autolysis loop south, exosite I east, and exosite II west. The slow form has residues 148-149e of the autolysis loop not defined in the electron density map. The remainder of the backbone trace is intact and documents no sites of autoproteolytic cleavage, although the structure was solved in the absence of inhibitors and crystal packing does not block the active site. The fast form was derived from the coordinates of the PPACK-inhibited thrombin (11), with the inhibitor removed for clarity.

Figure 3.
Fig. 3. A, environment of the catalytic triad in the slow form, shown with the 2F[o] F[c] electron density maps contoured at 1.0 level. The O- atom (B-factor = 27 Å2) of Ser-195 (B-factor = 27-33 Å2) rotates 54° and moves 1.2 Å relative to the position in the fast form inhibited with PPACK ( yellow). This change pertains to the slow form because it is not seen in the structure of the fast form inhibited with hirugen at exosite I and with the active site free (12). The shift in the position of Ser-195 breaks the H-bond with His-57. The interatomic distance between the O- of Ser-195 and the N- 2 (B-factor = 36 Å2) of His-57 (B-factor = 30-37 Å2) is 3.6 Å in the slow form, as opposed to 2.9 Å in the fast form. B, comparison of the active site cleft in the slow (colored by Corey-Pauling-Koltun) and fast form inhibited with PPACK (yellow, from PDB entry 1PPB). Hydrogen bonds are depicted as broken lines. Shown are the regions in contact with substrate (orange, shown with the P1-P3 residues) that are affected by the slow fast transition, i.e. the side chain of Asp-189 and Ser-195. Note how the shift in the position of Ser-195 breaks the H-bond with His-57, the imidazole group of which also rotates slightly relative to the position in the fast form. The O- atom of Ser-195 in the slow form moves too close to the C- atom of the P1 residue of substrate.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2002, 277, 40177-40180) copyright 2002.

Figures were selected by an automated process.