Conformational and rheological changes in catfish myosin as affected by different acids during acid-induced unfolding and refolding.

Changes in the conformation of catfish (Ictalurus punctatus) myosin due to (i) anions, (ii) acid pH, and (iii) salt addition were determined using tryptophan fluorescence, hydrophobicity measurements, differential scanning calorimetry, and circular dichroism. The relationship between conformation and storage modulus (G') of acid-treated myosin was studied. Three acids, HCl, H2SO4, and H3PO4, were used for unfolding myosin at three acidic pH conditions, 1.5, 2.0, and 2.5. Unfolded myosin was refolded to pH 7.3. Denaturation and unfolding of myosin was significantly (p < 0.05) lower when salt (0.6 M NaCl) was present during acid unfolding than in the absence of salt. When salt was added before unfolding, the alpha-helix content of myosin treated at pH 1.5 was significantly lower than that treated at pH 2.5. When salt was added after refolding, the alpha-helix content of myosin was unaffected by different pH treatments. The G' of myosin increased with an increase in myosin denaturation. The G' of myosin was significantly (p < 0.05) higher when salt was added to myosin after refolding than before acid unfolding. Among the different anion treatments, the G' of acid-treated myosin decreased in the order Cl- approximately SO42- > PO43-. Among the different pH treatments, the G' of myosin treated at pH 1.5 was significantly (p < 0.05) higher than myosin treated at pH 2.5. The conditions that would result in maximum myosin denaturation and maximum G' were unfolding of myosin at pH 1.5 using Cl- (from HCl) followed by refolding at pH 7.3 and subsequent addition of 0.6 M NaCl.