Abstract
The chaperonin system GroEL-GroES is present in all kingdoms of life and rescues proteins from improper folding and aggregation upon internal and external stress conditions, including high temperatures and pressures. Here, we set out to explore the thermo- and piezostability of GroEL, GroES and the GroEL-GroES complex in the presence of cosolvents, nucleotides and salts employing quantitative FTIR spectroscopy and small-angle X-ray scattering. Owing to its high biological relevance and lack of data, our focus was especially on the effect of pressure on the chaperonin system. The experimental results reveal that the GroEL-GroES complex is remarkably temperature stable with an unfolding temperature beyond 70 °C, which can still be slightly increased by compatible cosolutes like TMAO. Conversely, the pressure stability of GroEL and hence the GroEL-GroES complex is rather limited and much less than that of monomeric proteins. Whereas GroES is pressure stable up to ∼5 kbar, GroEl and the GroEl-GroES complex undergo minor structural changes already beyond 1 kbar, which can be attributed to a dissociation-induced conformational drift. Quite unexpectedly, no significant unfolding of GroEL is observed even up to 10 kbar, however, i.e., the subunits themselves are very pressure stable. As for the physiological relevance, the structural integrity of the chaperonin system is retained in a relatively narrow pressure range, from about 1 to 1000 bar, which is just the pressure range encountered by life on Earth.
Highlights
Chaperonins represent an important class of proteins in biological organisms, which have an essential role in assisting protein folding by transient encapsulation of nascent proteins in an ATP-driven mechanism to prevent misfolding or aggregation
In agreement with the marked changes observed in the p(r) function, these results indicate dissociation of the GroEL–GroES complex at pressures beyond about 2 kbar, which would be consistent with conclusions based on indirect measurements using light scattering methodology.[21]
The chaperonin system GroEL–GroES is present in all kingdoms of life and rescues proteins from improper folding and aggregation
Summary
Chaperonins represent an important class of proteins in biological organisms (archaea, bacteria and eukarya), which have an essential role in assisting protein folding by transient encapsulation of nascent proteins in an ATP-driven mechanism to prevent misfolding or aggregation. Stress-induced denaturation of proteins can be counteracted by the chaperonin’s support of correct refolding.[1,2,3,4] Here, we investigated the effect of temperature and pressure stress on the conformational stability of the heat shock protein (Hsp) complex GroEL– GroES of Escherichia coli (Fig. 1a), which belongs to the group I chaperonins. Chaperonins, like the GroEL–GroES complex, are oligomeric proteins, which consist of a large double-ring structure stacked back to back, thereby enclosing a central cavity with its lid-like cofactor. In this case, the lid-like cofactor refers to GroES, which contains seven identical B10 kDa subunits assembled as a heptamer ring and caps the ends of the GroEL cylinder. Two heptameric rings of GroEL (each subunit contains B57 kDa) form the cavity, exposing hydrophobic amino
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