Reviewer 1's Review

“Authors described interesting phenomenon in which nonspecific protein-protein interaction affects the formation and stability of triose phosphate isomerase, emphasizing the importance of the effect of the nonspecific protein-protein interaction on the specific protein-protein interaction. Taking into account that in the living cell specific protein-protein interactions are taking place in the presence of number of other proteins the effect of a nonspecific protein-protein interaction on a specific protein-protein interaction might be expected.

The hypothesis explaining the effect of nonspecific protein interaction on the refolding of triose phosphate isomerase was written on the basis of the literature supporting this hypothesis.

Authors followed the effect of several proteins on renaturation of triosephosphate isomerase denatured in guanidinium chloride. They observed the significant effect of the nonspecific interaction on the recovery and the velocity of renaturation process. They also investigated calorimetric effect of triosephosphate isomerase interaction with lysozyme. They concluded that the effect of the nonspecific protein-protein interaction on the renaturation of the isomerase depends on the interaction of protecting proteins with the active dimer of the isomerase. Quite convincing hypothesis, which could be even more convincing if the authors would determine the binding constants of the isomerase with protective proteins. Unspecific protein:protein interaction results in heterologous complex formation. In a such complex both proteins are in a more stabilized conformation. It is the reason that BSA, the extracellular protein, is commonly used to protect the cytosolic enzymes.

The best way to investigate the effect of the protective protein on the triose phosphate isomerase renaturation would be the determination of the binding constants of the complexes of protecting proteins with triose phosphate isomerase observed at the particular steps of this process, like Mu-BSA, Mf-BSA, Din-BSA and Dac-BSA. Having an adequate equipment it should be possible to perform such determination. But in any case it should be relatively easy to determine the binding constants of Dac with protective proteins. Determination of the binding constants would greatly improve the manuscript.

It is hard to overestimate the importance of protein-protein interaction in biology, in any case it is the interesting phenomenon for biochemists, biophysicist and molecular physiologists.

From protein chemists point of view it would be of interest to find the complementary domains of the protective protein like BSA and investigated protein like triose phosphate isomerase. For molecular physiologist it would be of interests to determine which unspecific protein-protein interactions are of importance for specific protein-protein interaction in vivo.”

N.B. These are the general comments made by the reviewer when reviewing this paper in light of which the manuscript was revised. Specific points addressed during revision of the paper are not shown.