liu.seSearch for publications in DiVA
Change search
ReferencesLink to record
Permanent link

Direct link
Mapping the Different Interactions between Eukaryotic β-actin and the Group I (GroEL) and Group II (TRiC) Chaperonins
Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
Department of Physics, The Norwegian University of Science and Technology, 7491 Trondheim, Norway.
Show others and affiliations
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Productive folding to the native state of the abundant eukaryotic protein actin is dependent on the chaperonin TRiC. The prokaryotic chaperonin GroEL also recognizes actin, but this interaction does not lead to the correct folding of actin. It is well established that GroEL interacts with non-native proteins through hydrophobic interactions. The characteristics of the interactions between TRiC and its target proteins are however unclear. In this study, we present multiple site-directed cysteine labeling and fluorescence measurements indicating that actin initially binds to TRiC through several interaction sites and that the surfaces of the interaction areas on the walls of the TRiC chamber present both polar and hydrophobic residues. At a later stage in the chaperonin cycle, the binding of ATP causes conformational changes in the chaperonin, which leads to a presentation of a more hydrophobic milieu in TRiC chamber. The conformational changes of the chaperonin causes rearrangements of the actin molecule and new interactions are proposed to be formed. Additionally, we show that the initial binding of actin to TRiC leads to a re-modeling of the nucleotide-binding cleft in actin. We also present data indicating that GroEL presents less specific interaction areas towards the bound actin than TRiC does. The interactions between actin and GroEL are tight and of hydrophobic character. No re-modeling of the nucleotide-binding cleft was obtained in the actin-GroEL complex. We conclude that the interactions between actin and TRiC are of both polar and hydrophobic character, the nature of the interactions are different in the prokaryotic and eukaryotic chaperonins, and the rearrangements of the nucleotide binding cleft of actin seen in the chaperonin cycle of TRiC do not occur in GroEL. We suggest that the rearrangements of the nucleotide-binding site in actin are critical for productive folding of actin.

National Category
Natural Sciences
URN: urn:nbn:se:liu:diva-52934OAI: diva2:285943
Available from: 2010-01-13 Created: 2010-01-13 Last updated: 2010-01-13
In thesis
1. Structural rearrangements of actins interacting with the Chaperonin systems TRiC/Prefoldin and GroEL/ES
Open this publication in new window or tab >>Structural rearrangements of actins interacting with the Chaperonin systems TRiC/Prefoldin and GroEL/ES
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The studies in this thesis are mainly focused on the effects that the chaperonin mechanisms have on a bound target protein. Earlier studies have shown that the bacterial chaperonin GroEL plays an active role in unfolding a target protein during the initial binding. Here, the effects of the eukaryotic chaperonin TRiC’s mechanical action on a bound target protein were studied by fluorescence resonance energy transfer (FRET) measurements by attaching the fluorophore fluorescein to specific positions in the structure of the target protein, β-actin. Actin is an abundant eukaryotic protein and is dependent on TRiC to reach its native state. It was found that at the initial binding to TRiC, the actin structure is stretched, particularly across the nucleotide-binding site. This finding led to the conclusion that the binding-induced unfolding mechanism is conserved through evolution. Further studies indicated that in a subsequent step of the chaperonin cycle, the actin molecule collapses. This collapse leads to rearrangements of the structure at the nucleotide-binding cleft, which is also narrowed as a consequence.

As a comparison to the productive folding of actin in the TRiC chaperonin system, FRET studies were also performed on actin interacting with GroEL. This is a non-productive interaction in terms of guiding actin to its native state. The study presents data indicating that the nucleotide-binding cleft in actin is not rearranged by GroEL in the same way as it is rearranged during the TRiC interaction. Thus, it could be concluded that although the general unfolding mechanism is conserved through the evolution of the chaperonins, an additional and specific binding to distinct parts of the actin molecule has evolved in TRiC. This specific binding leads to a directed unfolding and rearrangement of the nucleotide-binding cleft, which is vital for actin to reach its native state. The differences in the chemical properties of the actin-GroEL and the actin-TRiC complexes were also determined by measurements of fluorescein anisotropies and AEDANS emission shifts for probes attached to positions spread throughout the actin structure.

The evolutionary aspects of the chaperonin mechanisms and the target protein binding were further investigated in another study. In this study, the prokaryotic homologue to actin, MreB, was shown to bind to both TRiC and GroEL. MreB was also shown to bind to the co-chaperonin GroES.

In a separate study, the interaction between actin and the chaperone prefoldin was investigated. In vivo prefoldin interacts with non-native actin and transfers it to TRiC for subsequent and proper folding. In this homo-FRET study, it was shown that actin binds to prefoldin in a stretched conformation, similar to the initial binding of actin to TRiC.

Place, publisher, year, edition, pages
Institutionen för fysik, kemi och biologi, 2007. 72 p.
Linköping Studies in Science and Technology. Dissertations, ISSN 0345-7524 ; 1099
Biochemistry, chaperonin mechanisms, bound target protein, fluorescence resonance energy transfer (FRET), Actin, abundant eukaryotic protein
National Category
Biochemistry and Molecular Biology
urn:nbn:se:liu:diva-11445 (URN)978-91-85715-05-3 (ISBN)
Public defence
2007-05-25, Planck, Fysikhuset, Campus Valla, Linköpings universitet, Linköping, 14:00 (English)
On the day of the defence date the satus of article I was: In press.Available from: 2008-04-03 Created: 2008-04-03 Last updated: 2010-01-13Bibliographically approved

Open Access in DiVA

No full text

Search in DiVA

By author/editor
Villebeck, LailaKlang, HannaMoparthi, Satish BabuHammarström, PerJonsson, Bengt-Harald
By organisation
Molecular Biotechnology The Institute of TechnologyCell BiologyFaculty of Health SciencesDepartment of Physics, Chemistry and BiologyBiochemistry
Natural Sciences

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Total: 50 hits
ReferencesLink to record
Permanent link

Direct link