Comparative Fourier transform infrared and circular dichroism spectroscopic analysis of alpha(1)-proteinase inhibitor and ovalbumin in aqueous solution

alpha (1)-Proteinase inhibitor (alpha (1)Pi) and ovalbumin are both members of the serpin superfamily. They share about a 30% sequence identity and ex hibit great similarity in their three-dimensional structures. However, no a pparent functional relationship has been found between the two proteins. Un like alpha (1)Pi, ovalbumin shows no inhibitory effect to serine proteases. To see whether or not a conformational factor(s) may contribute to the fun ctional difference, we carried out comparative analysis of the two proteins ' secondary structure, thermal stability, and H-D exchange using FT-IR and CD spectroscopy. FT-IR analysis reveals significant differences in the amid e I spectral patterns of the two proteins. Upon thermal denaturation, both proteins exhibit a strong low-wavenumber beta -sheet band at 1624 cm(-1) an d a weak high-wavenumber beta -sheet band at 1694 cm(-1), indicative of int ermolecular aggregate formation. However, the midpoint of the thermal-induc ed transition of alpha (1)Pi (similar to 55 degreesC) is 18 degreesC lower than that of ovalbumin (similar to 73 degreesC). The thermal stability anal ysis provides new insight into the structural changes associated with denat uration. The result of H-D exchange explains some puzzling spectral differe nces between the two proteins in D2O reported previously. (C) 2000 Academic Press.