Circular dichroism and the secondary structure of the ROF2 protein from Arabidopsis thaliana

The protein ROF2 from the plant Arabidopsis thaliana acts as a heat stress modulator, being involved in the long-term acquired thermotolerance of the plant. Here we investigate the relationship between the biological function and the structure of ROF2, inferred by circular dichroism (CD) spectroscopy. The far-UV CD spectra, analyzed with the CDPro and DICHROWEB program packages, yield the percentages of alpha-helices, beta-sheets, unordered regions, turns and poly(Pro)II-helices in the secondary structure of ROF2. According to the analysis, the percentages of the structural elements of ROF2 are about 40% for beta-sheets, 30% for unordered regions, 17% for turns, 10% for poly(Pro)II-helices and 3% for alpha-helices. The near-UV CD spectra suggest that ROF2 proteins can associate, forming super-secondary structures. Our CD experiments performed at temperatures between 5 A degrees C and 97 A degrees C indicate that the thermal denaturation of ROF2 caused by a raise in temperature up to 55 A degrees C is followed by a thermal refolding of the protein as the temperature is raised further. The new secondary structure, acquired around 65 A degrees C, remains stable up to 97 A degrees C. The structural stability of ROF2 at high temperatures might play an important role in the experimentally observed thermotolerance of Arabidopsis thaliana.