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Stability and Folding of Proteins. Introduction

B. I. Kurganov

Bakh Institute of Biochemistry, Russian Academy of Sciences, Leninskii pr. 33, Moscow, 117071 Russia; fax: (095) 954-2732; E-mail: inbio@glas.apc.org

B. I. Kurganov
B. I. Kurganov

The present and following issues of Biochemistry (Moscow) are a continuation of a series of subject issues devoted to the key problems of modern biochemistry and related sciences. The topics of this issue involve the problems of stability and folding of proteins.
KEY WORDS: stability and folding of proteins

One of the important physicochemical characteristics of proteins is the degree of their resistance to various denaturing factors. The protein stability characteristics are used in practical work with proteins and in biotechnological industry. Investigation of the complete unfolding of a protein from an ordered native state (N) to a disordered unfolded state (U) can provide key thermodynamic information about the stability of a protein. Chief among these thermodynamic parameters is the free energy change for the N <--> U transition, which is a measure of the stability of the N state with respect to the U state.

Various physicochemical methods are applied for the study of thermodynamics and kinetics of protein denaturation processes and the structure of the unfolded states of proteins, among which are differential scanning calorimetry, circular dichroism, light scattering, X-ray, and neutron scattering, and fluorescent methods (the reviews by M. R. Eftink, G. Damaschun with coworkers, W. Pfeil, R. Jaenicke, and D. I. Levitsky with coworkers). One of the new and very informative methods of the study of protein stability is a protein fragmentation/mass spectrometry approach for determining amide hydrogen exchange rates in short segments of intact proteins following their incubation in D2O (D. L. Smith).

The investigations of denaturation of enzymes showed that the active sites are more sensitive to the action of denaturing factors than the entire protein molecule. As a result, the loss of enzymatic activity precedes the global unfolding of the protein molecule (C.-L. Tsou).

The special features of denaturation of oligomeric enzymes are due to the change in the quaternary structure of the protein under study in the course of the denaturation process. According to O. M. Poltorak and E. S. Chukhray, dissociative denaturation of oligomeric enzymes involves the stage of breakdown of "conformational lock" (the latter may be defined as the structural elements of the intersubunit contacts whose disruption makes possible the dissociation of a protein oligomer into more labile subunits). Recent advances in the development of the concept of conformational lock are connected with deciphering those peculiarities of the structure of intersubunit contacts which give an insight into the stepped character of their disruption during denaturation (alkaline phosphatase from Escherichia coli and horse liver alcohol dehydrogenase are used as examples).

The role of cofactors in stabilization of proteins and the effects of specific ligands on the stability of protein molecules have been reviewed by V. N. Uversky and N. V. Narizhneva.

The discussion of the stability of proteins in organic media (A. K. Gladilin and A. V. Levashov) and the methods of enzyme stabilization via chemical modification (R. Tyagi and M. N. Gupta) are of special interest in connection with the use of enzymes in biotechnological industry.

Until recently the mechanisms of loss of the biological activity of lyophilized proteins used in pharmacy and other applied fields remained poorly explored. The review by H. R. Costantino with colleagues fills this gap. The above-mentioned researchers showed that the proteins which have both disulfide bonds and free thiol groups are able to aggregate via thiol-disulfide exchange, and this process may be facilitated by lyophilization-induced structural perturbations.

Folding of polypeptide chain into the spatial structure possessing biological activity is one of the key problems in biochemistry and molecular biology. Until the present time the idea of C. B. Anfinsen declaring that amino acid sequence of polypeptide contains all information necessary for its folding in the native structure was prevailing. However, as experimental data accumulated it became clear that folding of proteins in the cell depends on auxiliary proteins, part of them being denominated as a group of molecular chaperons. The main function of chaperons recognizing nonfolded or partially denatured forms of proteins is preventing incorrect association of polypeptide chains leading to their aggregation. The mechanisms of the functioning of chaperons are discussed in the reviews by J. Martin, M. T. Fisher, and V. V. Mesyanzhinov with coworkers. Specific enzymes, peptidyl prolyl cis-trans isomerase and protein disulfide isomerase, are involved in protein folding. According to Ch.-ch. Wang, protein disulfide isomerase is not only an isomerase catalyzing the formation of native disulfide bonds in polypeptides being synthesized, but also a molecular chaperon participating in protein folding process. The review by O. B. Ptitsyn is devoted to modern hypotheses explaining the mechanism of folding of a polypeptide chain. Theoretical description of protein folding is based on the use of principles of mean force potentials and radial distribution functions as defined in statistical mechanics (W. A. Koppensteiner and M. J. Sippl).

The two-step procedure of in vitro refolding of proteins with the participation of artificial chaperons proposed by D. Rozema and S. H. Gellman has been inspired by the two-step mechanism of the functioning of GroE system. In the first step, the protein is captured by a detergent under conditions that would normally lead to irreversible protein aggregation. In the second step, removal of detergent from the protein--detergent complex is triggered by addition of a cyclodextrin which is capable of forming "inclusion complexes" with detergent, allowing the protein to refold. The examples of refolding of proteins with the participation of artificial chaperons are discussed in the review by B. I. Kurganov and I. N. Topchieva.

I hope that articles published in these issues will be of interest to biochemists and molecular biologists concerned with the fundamental problems of stability and folding of proteins as well as to biotechnologists elaborating new pharmaceutical protein preparations or applying protein (enzyme) preparations for synthesis of various compounds.

To facilitate access to these articles, the English versions of these review issues of Biochemistry (Moscow) will be made available on the World-Wide-Web at http://www.protein.bio.msu.su/biokhimiya long before their appearance in print.

Guest Editor of this issue,
Professor B. I. Kurganov
Bakh Institute of Biochemistry,
Russian Academy of Sciences,