Is the continuity of the domains required for the correct folding of a two-domain protein?


The role of domains in protein folding has been widely studied and discussed. Nevertheless, it is not clear whether the continuity of the domains in a protein is an essential requirement in determining the folding pathway. Previous studies have shown that the isolated structural domains of the two-domain monomeric enzyme, yeast phosphoglycerate kinase (yPGK), are able to fold independently into a quasinative structure, but they neither reassociate nor generate a functional enzyme [Minard, P., Hall, L., Betton, J. M., Missiakas, D., & Yon, J. M. (1989) Protein Eng. 3, 55-60; Fairbrother, W. J., Bowen, D., Hall, L., Williams, R. J. P. (1989) Eur. J. Biochem. 184, 617-625; Missiakas, D., Betton, J. M., Minard, P., & Yon, J. M. (1990) Biochemistry 29, 8683-8689]. In the present work, two circularly permuted variants of the yPGK gene were constructed. The natural adjacent chain termini were directly connected and the new extremities were created within the N-domain (at residues 71 and 72) or the C-domain (at residues 291 and 292), respectively. These two proteins were overexpressed and purified. They exhibit 14% and 23% of the wild-type enzyme activity, respectively. The two mutants fold in a compact conformation with slight changes in the secondary and tertiary structure probably related to the presence of a heterogeneous population of molecules. The unfolding studies reveal a large decrease in stability. From the present data it appears that, although the circular permutations induce some perturbations in the structure and stability of the protein, the continuity of the domains is not required for the protein to reach a native-like and functional structure. Study holds ProTherm entries: 4933 Extra Details: additive : EDTA(0.5 mM), domains; enzyme activity; secondary and tertiary structure;,circular permutations; functional tructure

Submission Details

ID: pBA7edTh3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:27 p.m.

Version: 1

Publication Details
Ritco-Vonsovici M;Minard P;Desmadril M;Yon JM,Biochemistry (1995) Is the continuity of the domains required for the correct folding of a two-domain protein? PMID:8527427
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