Binding of fibrinogen A alpha 1-50-beta-galactosidase fusion protein to thrombin stabilizes the slow form.
The interaction of fibrinogen A alpha1-50-beta-galactosidase fusion protein with the slow and fast forms of thrombin was studied and compared to thrombin-fibrinogen interaction under identical solution conditions. At equilibrium, the affinity of the fusion protein for the slow form of thrombin is 3 times higher than its affinity for the fast form. The fusion protein and fibrinogen have the same affinity for the fast form. On the other hand, the affinity of the fusion protein for the slow form of thrombin is 40 times tighter than that of fibrinogen. In the transition state, binding of the fusion protein has the same properties as fibrinogen, with the fast form showing higher specificity. The N-terminal fragment of the fibrinogen A alpha chain thus contains residues that are responsible for the preferential binding of the fusion protein to the slow form at equilibrium and to the fast form in the transition state. If this fragment binds to thrombin in a similar way for fibrinogen and the fusion protein, then the N-terminal domains of the B beta and gamma chains of fibrinogen, that are not present in the fusion protein, must play a key role in the binding of fibrinogen to thrombin at equilibrium. These chains may destabilize binding to the slow form by nearly 2.4 kcal/mol, thereby favoring binding of fibrinogen to the fast form. We propose that the three chains of fibrinogen play different roles in the thrombin-fibrinogen interaction, with the A alpha chain containing residues for preferential binding to the fast form in the transition state and the B beta and gamma chains containing residues that destabilize binding to the slow form at equilibrium.
Lord
S T
ST
Department of Pathology, University of North Carolina, Chapel Hill 27599-7525, USA.
Rooney
M M
MM
Hopfner
K P
KP
Di Cera
E
E
D004795:Enzyme Stability; D005340:Fibrinogen / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D013816:Thermodynamics; D013917:Thrombin / Q000737:chemistry / Q000378:metabolism*; D001616:beta-Galactosidase / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D005340:Fibrinogen; D001616:beta-Galactosidase; D013917:Thrombin