80 INFLUENCE OF TEMPERATURE ON BIOLOGICAL SYSTEMS 



forms of myosin according to light scattering studies (2, 18, 19, 24). The 

 equilibrium may also play a physiological role, since the native and de- 

 natured forms are almost equally abundant at pH 7. Its associated heat 

 of reaction may enter the heat budget of the muscle, and its ion sensitivity 

 may fit it for a role in the contractile process. 



SUMMARY 



Evidence is presented that reversible denaturation plays an important 

 role in the myosin ATP-ase system, as shown especially by the differences 

 in the effects of pressure at different pH. A native site of the enzyme 

 may split ATP, with a heat of activation of about 31,000 cal/mole and a 

 volume increase on activation of about 60 cc/mole. On the other hand, 

 the site may lose a hydrogen ion with a pK near 7 and become reversibly 

 denatured, with a heat of reaction of about 12,600 cal/mole and an increase 

 in volume of about 100 cc/mole. The pK and the heat suggest that re- 

 versible denaturation is accompanied by the dissociation of a hydrogen 

 ion from a histidine residue of the enzyme. The large volume increase, how- 

 ever, shows that this dissociation is accompanied by an unfolding of the 

 enzyme or a loss of bound water. 



If the pK is 7 and the heat of reaction is 31,000 cal/mole, the entropy 

 of reaction is +10.7 cal/degree-mole. The entropy change for dissociation 

 of an imidazole proton with a pK of 6.8 at 25°C and a heat of dissociation 

 of 7200 cal/mole is —7.1 cal/degree-mole. The corrected entropy change 

 for the protein is then +17.8 cal/degree-mole. This is a small change for 

 a protein molecule, and it indicates that the denatured molecule is not 

 much more disordered than the native. 



The analysis shows that the effect of pressure opposes those of tempera- 

 ture and hydroxyl ion in both reversible denaturation and the splitting of 

 the enzyme-substrate complex. That the three factors all increase the 

 measured ATP-ase activity at pH above 7.4 results from the interplay of 

 effects on the two processes. The late discovery of reversible denaturation 

 in the well-studied myosin ATP-ase system appears to be explained by 

 the small heat of denaturation, by the relatively high pressures needed to 

 show departure from a linear pressure relation, by the lack of earlier 

 studies of the pH dependence of the pressure effects, and perhaps by a 

 difference in behavior of the system in sodium as opposed by potassium 

 salt solutions. 



REFERENCES 



1. Bailey, K. Structural proteins II. Muscle. In: The Proteins (ed. by H. Neurath 

 and K. Bailey) II, B: 951-1055. New York: Acad. Press, 1954. 



2. Blum, J. J. The enzymatic interaction between myosin and nucleotides. Arch. 

 Biochem. and Biophys. 55: 486-511. 1955. 



