Calorimetric studies on monomeric and polymeric actin.

Differential scanning calorimetry of polymeric F-actin at pH 8.0 showed that the polymer had a concentration-independent thermal profile with a single transition temperature of 81 degrees C. In contrast, the thermal profile of G-actin was concentration-dependent, and although it resembled the F-actin profile at lower concentrations, it was found to have a more complex profile at higher protein concentrations.

Temperature-dependent changes in the activity and tryptic susceptibility of membrane-bound transhydrogenase.

Beef heart mitochondrial pyridine nucleotide transhydrogenase undergoes a 2-fold changes in apparent activation energy between 5 degrees and 40 degrees C, from 30 kcal/mol at the lower end to 15 kcal/mol at the upper end of this range. In order to determine whether changes in the structural domain of transhydrogenase accompany the large change in activation energy, the degree of proteolytic inactivation of membrane-bound transhydrogenase was monitored as a function of temperature. Changes in the susceptibility of transhydrogenase to proteolytic inactivation over the 5 degrees -40 degrees C interval correlate well with the observed alteration in activation energies. No temperature-dependent change in either apparent activation energy or susceptibility to proteolysis were observed in detergent-solubilized transhydrogenase. Interactions between transhydrogenase and its microenvironment in the mitochondrial membrane may be an important factor in determining its activity.

Calorimetric studies of lipid phase transitions in native and heat-denatured membranes of beef heart submitochondrial particles.

The lipids in beef heart submitochondrial particles undergo a broad reversible endothermic phase change centered at about -10 degrees C. Following protein denaturation, a new reversible transition centered at about 20 degrees C appears. The extracted lipids from these membranes exhibit thermal behavior that is essentially identical to the lipid transition in the intact membrane after protein denaturation. A role for this latent pool of higher-melting lipids is proposed.

Effects of substrate and inhibitor binding on thermal and proteolytic inactivation of rat liver transhydrogenase.

The thermostability and proteolytic inactivation of rat liver submitochondrial particle transhydrogenase was studied in the presence of pyridine dinucleotide substrates and a variety of divalent metal and nucleotide inhibitors. Relative to the unliganded enzyme, the NADPH-enzyme complex was more thermostable and showed a twofold greater rate of tryptic inactivation, while the NADP+-enzyme complex was more thermolabile and only slightly more susceptible to tryptic inactivation. Neither NAD+ nor NADH significantly affected thermostability or proteolysis. Similar effects of these ligands were observed for the non-energy-linked and energy-linked transhydrogenase reactions, indicating that both activities are catalyzed by the same enzyme. In thermal experiments, acetyl-CoA, 2'-AMP, and NMNH stabilized, palmitoyl-CoAlabilized, and dephospho-CoA, CoA, NMN+, and 5'-AMP had little effect on enzyme stability. Tryptic inactivation was inhibited by 2'-AMP and NMN+ but was not influenced by the other nucleotide inhibitors. Divalent metal ion inhibitors (Mg2+, Ca2+, Mn2+, Ba2+, and Sr2+) stabilized transhydrogenase against thermal inactivation and promoted tryptic inactivation.