The unfolded ß-barrel of SOD1 is in a compact state, stabilised by long-range hydrophobic contacts.

• The unfolded state of a globular protein in a physiologically relevant environment is by no means an inert random coil.  On the contrary, its structural and dynamic properties are crucial for e.g., protein folding and aggregation.  Despite its importance, it has been studied relatively sparsely, which is partly due to its low population which tend to obstruct detailed biophysical characterization.  Here, introduction of two destabilizing core mutations allow us to study the unfolded state of the central b-barrel of Superoxide Dismutase 1 under native conditions.  In order to structurally characterise the unfolded state, we use high-resolution nuclear magnetic resonance (NMR), including paramagnetic relaxation enhancement, to obtain constraints for the generation of unfolded ensembles.  The results show that the unfolded state is more compact than the chemically denatured state of the same protein.  This compacted state seems to be stabilised by long-range hydrophobic contacts, out of which many coincide with those found in the native state.  We also investigated the previously observed destabilising effect on the unfolded state by a poly-anion, and find that; the interaction does not alter the overall ensemble dimensions, nor the pattern in native-like contacts.  On the other hand, addition of the chemical denaturant urea results in a more expanded state.  The varying compaction with different co-solutes was validated by pulsed-field gradient NMR diffusion measurements.  Unlike helical proteins, b-proteins lack the ability to fulfil hydrogen bonds by local native interactions. This forces specific prerequisites on the collapsed pre-folding state.  Here, the compaction is enabled by both native-like and non-native long-range contacts in the unfolded ensemble, and we suggest that the average topology of the collapsed state is determined by the sequence distribution of hydrophobic patches, separated by non-interacting hydrophilic clusters. 

Ämnesord

NATURVETENSKAP  -- Biologi -- Biofysik (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Biophysics (hsv//eng)

NATURVETENSKAP  -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)

Nyckelord

biokemi

Biochemistry

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vet (ämneskategori)

ovr (ämneskategori)