DOMAIN-STRUCTURE OF THE FIB-1 AND FIB-2 REGIONS OF HUMAN FIBRONECTIN - THERMODYNAMIC PROPERTIES OF THE TYPE-I FINGER MODULE

The stability of the Fib-1 (29 kDa) and Fib-2 (19 kDa) fragments of human fibronectin as well as several different subfragments and isolated type I “finger” modules were studied under various solvent conditions by differential scanning calorimetry and fluorescence spectroscopy. It was established that all fibronectin fingers constitute independently folded domains whose melting temperatures range from 54 to 108°C. The difference between heat capacities of the native and denatured states (ΔC(p)) is low, about 0.03 cal/K-g, which is consistent with the relatively low percentage of hydrophobic amino acids and the consequent small change in non-polar surface area exposed to the solvent upon denaturation. The free energy of unfolding at 25°C, as calculated from the calorimetric data or measured directly by titration with GdmSCN is also small, in the range of 2.4 to 6.7 kcal/mol. The small ΔG value and its flat dependence on temperature (determined by ΔC(p)) translates the small variations in ΔG between fingers into large variations in t(m). The small value of ΔG also indicates that finger modules are structurally rather fragile which may account for their sensitivity to proteolysis; almost any cleavage within either of the two disulfide loops destroys the cooperative structure and abolishes the corresponding melting transition. The fact that some fingers exhibit large decreases in t, l, upon separation from more stable neighbors with which they interact can also be viewed as a consequence of the low values of ΔG and ΔG(p). © 1994 Academic Press Limited.