Domain behavior during the folding of a thermostable phosphoglycerate kinase.


Abstract

Bacillus stearothermophilus phosphoglycerate kinase (bsPGK) is a monomeric enzyme of 394 residues comprising two globular domains (N and C), covalently linked by an interdomain alpha-helix (residues 170-185). The molecule folds to the native state in three stages. In the first, each domain rapidly and independently collapses to form an intermediate in which the N-domain is stabilized by 5.1 kcal mol-1 and the C-domain by 3.3 kcal mol-1 over their respective unfolded conformations. The N-domain then converts to a folded state at a rate of 1.2 s-1 (delta GI-F = 3.8 kcal mol-1), followed by the C-domain at 0.032 s-1 (delta GI-F = 12.1 kcal mol-1). It is this last step that limits the rate of acquisition of enzyme activity. In the dynamics of unfolding in water, the N-domain converts to the intermediate state at a rate of 8 x 10(-4) s-1, some 10(7) times faster than the C-domain. Consequently, the most populated intermediate in the folding reaction has a native-like N-domain, while that in the unfolding direction has a native-like C-domain. In a conventional sense, therefore, the folding/unfolding kinetics of bsPGK can be described as random order. Consistent with these observations, cutting the molecule in the interdomain helix produces two, independently stable units comprising residues 1-175 and 180-394. A detailed comparison of their folding behavior with that of the whole molecule reveals that true interdomain contacts are relatively weak, contributing approximately 1.4 kcal mol-1 to the stability of the active enzyme. The only interactions which contribute to the stability of rapidly formed intermediates or to transition states along the productive folding pathways are those within domain cores. Contacts formed either between domains or with the interdomain helix are made only in the folded ground state, but do not constitute a separate step in the folding mechanism. Intriguingly, the most pronounced effect of interdomain contacts on the kinetics of folding is inhibitory; the presence of the C-domain appearing to reduce the effective rate of acquisition of native structure within the N-domain. Study holds ProTherm entries: 5339, 5340, 5341, 5342 Extra Details: globular domain; interdomain alpha-helix; dynamics of unfolding;,intermediate; transition states

Submission Details

ID: y5jD5JtV

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:29 p.m.

Version: 1

Publication Details
Parker MJ;Spencer J;Jackson GS;Burston SG;Hosszu LL;Craven CJ;Waltho JP;Clarke AR,Biochemistry (1996) Domain behavior during the folding of a thermostable phosphoglycerate kinase. PMID:8961937
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1PHP 1994-06-22 1.65 STRUCTURE OF THE ADP COMPLEX OF THE 3-PHOSPHOGLYCERATE KINASE FROM BACILLUS STEAROTHERMOPHILUS AT 1.65 ANGSTROMS

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
90.1 Phosphoglycerate kinase C5D7M4 PGK_GEOSW
98.2 Phosphoglycerate kinase Q5KVE4 PGK_GEOKA
100.0 Phosphoglycerate kinase P18912 PGK_GEOSE