Insights in the molecular steps of the coagulation cascade / Relevance for cell-membrane interactions
Researchers at the Max Planck Institute for Biochemistry in Martinsried/Munich have determined the structure of an important domain of coagulation factor Va. This opens new insights into the molecular details of blood coagulation, as well as in the overall mechanisms of protein-membrane association. The results are reported in Nature 25 November 1999.
An intact circulatory system is vital for all physiological processes. Vascular injuries must be therefore rapidly sealed and repaired. This is achieved through the so-called blood coagulation cascade: several, similarly assembled proteinases (i.e., enzymes, that cleave other proteins) are stepwise specifically activated, in a cumulative domino effect. In the final step of this coagulation cascade, the proteinase thrombin is liberated into the blood stream. As a result of thrombin’s activities, the soluble plasma protein fibrinogen is converted into fibrin, which spontaneously associates into insoluble polymers. Further, blood platelets are activated by thrombin, and aggregate thereupon. Fibrin and activated platelets constitute the main components of the blood clot, which seals the site of the injury.
A research group, headed by Wolfram Bode in the Department of Structural Research (Director: Prof. Robert Huber) at the Max Planck Institute for Biochemistry in Martinsried/Germany, has solved in the last decade the X-ray structures of thrombin and several inhibitor complexes, as well as other coagulation proteinases (factors IXa and Xa). These results have improved our understanding of the molecular processes leading to blood clotting. On the other hand, the structures of the non-enzymatic cofactors Va and VIIIa were up to now elusive, and therefore their mechanism of membrane association only poorly understood. Investigations conducted at the Duke University (Durham/North Carolina) have shown that the C-terminal domain of factor Va is essential for membrane attachment. This process is highly stereospecific, i.e. it relies on the recognition of phospholipid head groups that possess the amino acid L-serine, not its mirror image.
Pablo Fuentes-Prior and Sandra Macedo-Ribeiro of Wofram Bode’s group have solved, in collaboration with William Kane (Duke University), the crystal structures of this membrane binding domain of coagulation factor Va in two different crystal forms (see Figure). The now unraveled domain possesses a barrel-like shape, from which three major loops protrude. These loops either cover ("closed form") or disclose ("open form") a cavity suitable for accommodation of a negatively charged phospholipid head group. The researchers propose that interaction with a few phospholipid head groups in the "closed form" promotes conformational changes leading to the unfolding of the loops and the exposure of this major cavity. The loops can then insert into the interior of the membrane, helped by favorable electrostatic interactions between the negatively charged phospholipid head groups and basic amino acid residues located at the barrel surface.
The results of this structural analysis are relevant not only for the mechanism of blood clotting, but also for several other proteins involved in cell-cell and cell-membrane interactions. Only to mention one example: there are neuroreceptors (neuropilins), which possess similar domains and regulate the axon growth. Further, it is known that mutations in this functional unit are responsible for genetic diseases such as the X-linked juvenile retinoschisis. The determination of such a crucial structure constitutes therefore the basis for further investigations, in particular for mutagenesis studies.
Published: 25-11-99
Contact: Pablo Fuentes-Prior
Max Planck Institute for Biochemistry,
Martinsried/Germany
E-Mail: fuentes@biochem.mpg.de
Phone: (+49 89) 8578-2729
Fax: (+49 89) 8578-3516
Journal
Nature