Distribution of Charges inBacillus intermedins 7PRibonuclease Determines the Number of Cooperatively Melting Regions of the Globule

Abstract

A correlation between the distribution of charged side groups in the globule of Bacillus intermedius 7P ribonuclease (binase) and the process of heat denaturation was studied at different pH values in order to estimate a relation between charge distribution in globular proteins and the character of cooperative thermodynamic transitions. As was shown by comparing the results of scanning microcalorimetric analysis of heat denaturation with the three-dimensional structure of binase, at optimal pH the molecule exists as a single cooperative system stabilized by hydrogen bonds, Van der Waals' contacts, and electrostatic interactions like salt bridges. At pH lower than 4.0 (below the physiological optimum) the cooperativity type of the system was found to change due to a reversible cooperative transition in the ternary structure of the protein globule. It has been concluded that the molecular architecture and the arrangement of atoms do not change considerably in different environments; thus the thermodynamic properties of the globule vary due to the alteration of charge distribution and the consequent changes in the size and number of cooperative regions of the globule. Thus, structural and energetic domains may be non-coincident in proteins.

Heat denaturation of pepsinogen in a water‐ethanol mixture

“…Hence, we can conclude that the number of energetical domains in pepsinogen (as in pepsin [6]) does not change upon ethanol addition. However, as was shown in [2,6,14], the number of energetical domains may be changed following a pH shift• A similar conclusion can be drawn for pepsinogen (Table 1). An increase in pH of the aqueous solution results in an R decrease from 1.9 at pH 6.0 to 1.5, at pH 8.0.…”

Representation of amino acid sequences in terms of interaction energy in protein globules

“…These data agree with the fact that more than half of the substitutions are in regions 16–20 and 28–40. As to the data for binase and barnase presented in Table 1, we may conclude that the delineated segments show a striking correspondence to cooperative units defined in microcalorimetric experiments [20]. Besides, our results are in general accordance with the modular structure (1–24, 25–52, 53–73, 74–88, 89–98, and 99–110) revealed on the basis of the so‐called centripetal profile approach [21]and studied in biochemical experiments [22, 23].…”

The studies of cooperative regions in T7 RNA polymerase

“…This allows us to draw certain conclusions about the sizes of the independently melting units of the T7RNAP molecule. As was shown in [7,15], the ratio of denaturation enthalpies of independently melting domains at 110°C is equal to the ra- [3,4] and from X-ray analysis [5] of T7RNAP. Fig.…”

“…The results obtained suggest that the T7RNAP molecule consists of two relatively independently melting cooperative units ('energetic domains' of our definition [7,8]) the sizes of which coincide with those of structural domains [5]. The role of the N-terminal domain in the stabilization of the C-terminal one and the whole enzyme structure is also discussed.…”