3P70: Structural Basis Of Thrombin-Mediated Factor V Activation: Essential Role Of The Hirudin-Like Sequence Glu666-Glu672 For Processing At The Heavy Chain-B Domain Junction

Citation:
Abstract
Thrombin-catalyzed activation of coagulation factor V (FV) is an essential positive feedback reaction within the blood clotting system. Efficient processing at the N- (Arg(709)-Ser(710)) and C-terminal activation cleavage sites (Arg(1545)-Ser(1546)) requires initial substrate interactions with 2 clusters of positively charged residues on the proteinase surface, exosites I and II. We addressed the mechanism of activation of human factor V (FV) using peptides that cover the entire acidic regions preceding these cleavage sites, FV (657-709)/ (FVa2) and FV(1481-1545)/(FVa3). FVa2 appears to interact mostly with exosite I, while both exosites are involved in interactions with the C-terminal linker. The 1.7-A crystal structure of irreversibly inhibited thrombin bound to FVa2 unambiguously reveals docking of FV residues Glu(666)-Glu(672) to exosite I. These findings were confirmed in a second, medium-resolution structure of FVa2 bound to the benzamidine-inhibited proteinase. Our results suggest that the acidic A2-B domain linker is involved in major interactions with thrombin during cofactor activation, with its more N-terminal hirudin-like sequence playing a critical role. Modeling experiments indicate that FVa2, and likely also FVa3, wrap around thrombin in productive thrombin.FV complexes that cover a large surface of the activator to engage the active site.
PDB ID: 3P70Download
MMDB ID: 93874
PDB Deposition Date: 2010/10/11
Updated in MMDB: 2011/09
Experimental Method:
x-ray diffraction
Resolution: 2.55  Å
Source Organism:
Similar Structures:
Biological Unit for 3P70: trimeric; determined by author and by software (PISA)
Molecular Components in 3P70
Label Count Molecule
Proteins (3 molecules)
1
Human Alpha-thrombin, Light Chain(Gene symbol: F2)
Molecule annotation
1
Human Alpha-thrombin, Heavy Chain(Gene symbol: F2)
Molecule annotation
1
Human Factor V, A2-b Domain Linker(Gene symbol: F5)
Molecule annotation
Chemicals (5 molecules)
1
1
2
1
3
2
4
1
* Click molecule labels to explore molecular sequence information.

Citing MMDB
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