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Reconstitution of collagen fibrils in vitro; the assembly process depends on the initiating procedure

1986; Elsevier BV; Volume: 8; Issue: 3 Linguagem: Inglês

10.1016/0141-8130(86)90020-6

1879-0003

David Holmes, Michael J. Capaldi, John A. Chapman,

Hydrogels: synthesis, properties, applications

Electron microscopy and turbidimetry have been used to show that the reconstitution of native-type banded fibrils from a solution of acetic acid-soluble collagen (with intact telepeptides) can proceed by different assembly pathways, depending on the initiating procedure used to bring the collagen from a dissolved to a precipitating condition. Initiation requires raising the pH, temperature (T) and ionic strength (I) of the solution (here from pH 3.4, T=4°C, I=0.01 to pH 7.4, T=34°C, I=0.2). A widely used procedure is an initial increase in pH and I, followed by a rise in T (the 'neutral start' procedure); this leads, in the early stages, to the mesh of long thin non-banded filaments described by Gelman et al.9. When, however, the initiation steps are performed in reverse order, first raising T and then increasing pH and I (the 'warm start' procedure), non-banded filaments are not found as an abundant long-lived intermediate in the assembly process: instead, native-type banded fibrils constitute most of the precipitated material from the earliest stages onwards, and fibril formation takes place more rapidly. The simultaneous raising of T, pH and I (in a third procedure) not only yields banded fibrils throughout the course of precipitation (with no accumulation of filaments) but also results in a further increase in the rate of fibril formation. Filament formation appears therefore to be triggered by exposure to cold neutral solvent during initiation. When exposure to cold neutral solvent is avoided, an alternative and more efficient mode of assembly, involving the simultaneous lateral and axial growth of D-periodic fibrils, occurs.