Regular Article
The Sequence of Cleavage of Fibrinopeptides from Fibrinogen is Important for Protofibril Formation and Enhancement of Lateral Aggregation in Fibrin Clots

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Abstract

Thrombin cleaves A fibrinopeptides from fibrinogen faster than B; cleavage of the B fibrinopeptides has been associated with enhanced lateral aggregation in the fibrin clot. We show that protofibrils from soluble desA fibrin examined by electron microscopy appear to be more loosely organized than those from desAB fibrin. Protofibril lengths were measured from the micrographs; histograms of the distribution of protofibril sizes at different points in the lag period are different for the two species and indicate that the associations leading to oligomer formation are not random. A kinetic model for oligomer formation was developed to investigate the implications of these histograms for protofibril assembly; the model suggests that oligomer-oligomer reactions are important and that some reactions occur preferentially. The overall appearance of negatively contrasted clots made from fibrin monomer missing both fibrinopeptides was similar to that of clots from molecules missing only the A fibrinopeptide, in contrast to the large differences in clots produced by the action of the enzymes on fibrinogen. However, examination of the band patterns by electron microscopy reveals that fibers from desAB fibrin are less well ordered than those from desA fibrin and often aperiodic. In agreement with the electron microscope results, maximum turbidity values are similar for both types of clots, although the initial turbidity rise is more rapid for desAB soluble fibrin clots. If thrombin is added to desA soluble fibrin during the lag period, there is increased lateral aggregation of fibers observed by electron microscopy and the maximum turbidity is higher. These results indicate that there must be a delay in the cleavage of the B fibrinopeptides for the enhancement of lateral aggregation. Changes that occur upon fibrinopeptide B cleavage from molecules in a fiber, such as an increase in calcium ion binding or conformational changes, cause fibers to aggregate with each other or additional protofibrils to be added to a fiber. On the other hand, if these differences are present initially, as they are on soluble desAB fibrin, these results indicate that all steps occur more rapidly, producing relatively thin fibers that are not well ordered. In conclusion, the usual delay in fibrinopeptide B cleavage appears to be necessary both for normal protofibril and fiber assembly, so that the changes that accompany removal of these peptides preferentially affect lateral aggregation rather than earlier steps of fibrin clot assembly.

Keywords

fibrinogen
fibrinopeptides
lateral aggregation
protofibrils
polymerization kinetics

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