Compaction of ribosomal protein S6 by sucrose occurs only under native conditions.
The effect of osmolyte sucrose on the stability and compaction of the folded and unfolded states of ribosomal protein S6 from Thermus thermophilus was analyzed. Confirming previous results obtained with sodium sulfate and trehalose, refolding stopped-flow measurements of S6 show that sucrose favors the conversion of the unfolded state ensemble to a highly compact structure (75% as compact as the folded state). This conversion occurs when the unfolded state is suddenly placed under native conditions and the compact state accumulates in a transient off-folding pathway. This effect of sucrose on the compaction of the unfolded state ensemble is counteracted by guanidinium hydrochloride. The compact state does not accumulate at higher guanidinium concentrations and the unfolded state ensemble does not display increased compaction in the presence of 6 M guanidinium as evaluated by collisional quenching of tryptophan fluorescence. In contrast, accessibility of the tryptophan residue of folded S6 above 1 M sucrose concentration decreased as a result of an increased compaction of the folded state. Unfolding stopped-flow measurements of S6 reflect this increased compaction of the folded state, but the unfolding pathway is not affected by sucrose. Compaction of folded and unfolded S6 induced by sucrose occurs under native conditions indicating that decreased protein conformational entropy significantly contributes to the mechanism of protein stabilization by osmolytes.
Centro de Engenharia Biológica e Química, Instituto Superior Técnico, Av. Rovisco Pais 1049-001, Lisboa, Portugal.
LuYang
L
Chen
José A B
JA
Ferreira
Sílvia M B
SM
Costa
Gonçalo J M
GJ
Cabrita
Daniel E
DE
Otzen
Eduardo Pinho
EP
Melo
D019791:Guanidine / Q000494:pharmacology; D007700:Kinetics; D011489:Protein Denaturation; D017510:Protein Folding; D038601:Ribosomal Protein S6 / Q000737:chemistry* / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D013395:Sucrose / Q000494:pharmacology*; D013816:Thermodynamics; D016963:Thermus thermophilus / Q000737:chemistry
D038601:Ribosomal Protein S6; D013395:Sucrose; D019791:Guanidine