Credit: Darren Baker
In recreating the structure of prothrombin, researchers have improved their understanding of how the clotting factor functions.
By deleting a disordered linker region, they were able to visualize the complete structure of prothrombin.
This deleted version was activated to thrombin much faster than the intact version of prothrombin.
And results suggested that cofactor Va enhances the activation of prothrombin by altering the architecture of the linker.
Enrico Di Cera, MD, of Saint Louis University in Missouri, and his colleagues reported these findings in PNAS.
Last year, Dr Di Cera’s team published the first structure of prothrombin. This structure lacked a domain responsible for interaction with membranes, and certain other sections were not detected by X-ray analysis.
Though the researchers were able to crystallize the protein, there were disordered regions in the structure they could not see.
Within prothrombin, there are 2 kringle domains connected by a linker region that intrigued the researchers because of its intrinsic disorder.
“We deleted this linker, and crystals grew in a few days instead of months, revealing, for the first time, the full architecture of prothrombin,” Dr Di Cera said.
The crystal structure revealed a contorted conformation where the domains are not vertically stacked, kringle-1 comes close to the protease domain, and the Gla-domain contacts kringle-2.
The researchers also found the deleted version of prothrombin is activated to thrombin much faster than intact prothrombin.
Specifically, deletion of the linker reduced the enhancement of thrombin generation by cofactor Va from the more than 3000-fold observed with wild-type prothrombin to 60-fold. So it appears that deletion of the linker mimics the effect of cofactor Va on prothrombin activation.
“It took us almost 2 years to discover that the disordered linker was the key,” Dr Di Cera said. “Finally, prothrombin revealed its secrets, and, with that, the molecular mechanism of a key reaction of blood clotting finally becomes amenable to rational drug design for therapeutic intervention.”
