3SQH: Crystal Structure Of Prethrombin-2 Mutant S195a In The The Open Form

Citation:
Abstract
Prethrombin-2 is the immediate zymogen precursor of the clotting enzyme thrombin, which is generated upon cleavage at R15 and separation of the A chain and catalytic B chain. The X-ray structure of prethrombin-2 determined in the free form at 1.9 A resolution shows the 215-217 segment collapsed into the active site and occluding 49% of the volume available for substrate binding. Remarkably, some of the crystals harvested from the same crystallization well, under identical solution conditions, diffract to 2.2 A resolution in the same space group but produce a structure in which the 215-217 segment moves >5 A and occludes 24% of the volume available for substrate binding. The two alternative conformations of prethrombin-2 have the side chain of W215 relocating >9 A within the active site and are relevant to the allosteric E*-E equilibrium of the mature enzyme. Another unanticipated feature of prethrombin-2 bears on the mechanism of prothrombin activation. R15 is found buried within the protein in ionic interactions with E14e, D14l, and E18, thereby making its exposure to solvent necessary for proteolytic attack and conversion to thrombin. On the basis of this structural observation, we constructed the E14eA/D14lA/E18A triple mutant to reduce the level of electrostatic coupling with R15 and promote zymogen activation. The mutation causes prethrombin-2 to spontaneously convert to thrombin, without the need for the snake venom ecarin or the physiological prothrombinase complex.
PDB ID: 3SQHDownload
MMDB ID: 95370
PDB Deposition Date: 2011/7/5
Updated in MMDB: 2011/12
Experimental Method:
x-ray diffraction
Resolution: 2.2  Å
Source Organism:
Similar Structures:
Biological Unit for 3SQH: monomeric; determined by author and by software (PISA)
Molecular Components in 3SQH
Label Count Molecule
Protein (1 molecule)
1
Thrombin Light Chain, Heavy Chain(Gene symbol: F2)
Molecule annotation
* Click molecule labels to explore molecular sequence information.

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