The Molten Globule, and Two-State vs. Non-Two-State Folding of Globular Proteins

Kuwajima, Kunihiro

doi:10.20944/preprints202002.0290.v1

Cited by 12 publications

(6 citation statements)

References 92 publications

(144 reference statements)

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“…The SAXS analysis ( Figure 11 ) also supports the presence of a non-native folding state for rRop as it suggests a less dense packing of the molecule in comparison to its parent protein, and a highly flexible, molten-globule state. Generally, molten globules are characterized [ 45 ] by the presence of some native-like secondary structure, and a lower compactness of the overall structure of the molecule, properties which agree with the characteristics of the folding state of rRop.…”

Section: Discussionmentioning

confidence: 99%

Probing Protein Folding with Sequence-Reversed α-Helical Bundles

Kefala

Amprazi

Mylonas

et al. 2021

IJMS

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Recurrent protein folding motifs include various types of helical bundles formed by α-helices that supercoil around each other. While specific patterns of amino acid residues (heptad repeats) characterize the highly versatile folding motif of four-α-helical bundles, the significance of the polypeptide chain directionality is not sufficiently understood, although it determines sequence patterns, helical dipoles, and other parameters for the folding and oligomerization processes of bundles. To investigate directionality aspects in sequence-structure relationships, we reversed the amino acid sequences of two well-characterized, highly regular four-α-helical bundle proteins and studied the folding, oligomerization, and structural properties of the retro-proteins, using Circular Dichroism Spectroscopy (CD), Size Exclusion Chromatography combined with Multi-Angle Laser Light Scattering (SEC-MALS), and Small Angle X-Ray Scattering (SAXS). The comparison of the parent proteins with their retro-counterparts reveals that while the α-helical character of the parents is affected to varying degrees by sequence reversal, the folding states, oligomerization propensities, structural stabilities, and shapes of the new molecules strongly depend on the characteristics of the heptad repeat patterns. The highest similarities between parent and retro-proteins are associated with the presence of uninterrupted heptad patterns in helical bundles sequences.

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Section: Discussionmentioning

confidence: 99%

Probing Protein Folding with Sequence-Reversed α-Helical Bundles

Kefala

Amprazi

Mylonas

et al. 2021

IJMS

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“…The broad distribution of protection against exchange in I bp ( Fig. 2 ), in the context of the narrower distributions for N, N′, I a , and U, implies an alternative structural ensemble, possibly a molten globule stabilized by the hydrophobic effect ( 26 , 27 ). Although aggregation precluded direct measurement of protection in I bp , the remarkable resistance to exchange for β4α4 in the presence of high concentrations of GdnHCl ( Figs.…”

Section: Discussionmentioning

confidence: 99%

A conserved folding nucleus sculpts the free energy landscape of bacterial and archaeal orthologs from a divergent TIM barrel family

Jain

Muneeruddin

Anderson

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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The amino acid sequences of proteins have evolved over billions of years, preserving their structures and functions while responding to evolutionary forces. Are there conserved sequence and structural elements that preserve the protein folding mechanisms? The functionally diverse and ancient (βα)1–8 TIM barrel motif may answer this question. We mapped the complex six-state folding free energy surface of a ∼3.6 billion y old, bacterial indole-3-glycerol phosphate synthase (IGPS) TIM barrel enzyme by equilibrium and kinetic hydrogen–deuterium exchange mass spectrometry (HDX-MS). HDX-MS on the intact protein reported exchange in the native basin and the presence of two thermodynamically distinct on- and off-pathway intermediates in slow but dynamic equilibrium with each other. Proteolysis revealed protection in a small (α1β2) and a large cluster (β5α5β6α6β7) and that these clusters form cores of stability in Ia and Ibp. The strongest protection in both states resides in β4α4 with the highest density of branched aliphatic side chain contacts in the folded structure. Similar correlations were observed previously for an evolutionarily distinct archaeal IGPS, emphasizing a key role for hydrophobicity in stabilizing common high-energy folding intermediates. A bioinformatics analysis of IGPS sequences from the three superkingdoms revealed an exceedingly high hydrophobicity and surprising α-helix propensity for β4, preceded by a highly conserved βα-hairpin clamp that links β3 and β4. The conservation of the folding mechanisms for archaeal and bacterial IGPS proteins reflects the conservation of key elements of sequence and structure that first appeared in the last universal common ancestor of these ancient proteins.

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“…Our aim was a test if such inclusions would lead to similar intra- and inter-molecular Ar-Ar interactions observed in the abovementioned analogous systems. Such motifs and interactions in the cores of CC structures could result in desirable properties such as the inclusion of chromophore and fluorophore probes for biophysical studies; 31 platforms for binding small molecules; 32 the introduction of sites for post-translational modification; 33 catalytic centers; 34 and sub-structures that promote electron transfer. 35…”

Section: Figurementioning

confidence: 99%

How coiled-coil assemblies accommodate multiple aromatic residues

Rhys

Dawson

Beesley

et al. 2021

Preprint

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Rational protein design requires understanding the contribution of each amino acid to a targeted protein fold. For a subset of protein structures, namely the α-helical coiled coils (CCs), knowledge is sufficiently advanced to allow the rational de novo design of many structures, including entirely new protein folds. However, current CC design rules center on using aliphatic hydrophobic residues predominantly to drive the folding and assembly of amphipathic α helices. The consequences of using aromatic residues--which would be useful for introducing structural probes, and binding and catalytic functionalities--into these interfaces is not understood. There are specific examples of de-signed CCs containing such aromatic residues, e.g., phenylalanine-rich sequences, and the use of polar aromatic resi-dues to make buried hydrogen-bond networks. However, it is not known generally if sequences rich in tyrosine can form CCs, or what CC assemblies these would lead to. Here we explore tyrosine-rich sequences in a general CC-forming background and resolve new CC structures. In one of these, an antiparallel tetramer, the tyrosine residues are solvent accessible and pack at the interface between the core and the surface. In the other more-complex structure, the residues are buried and form an extended hydrogen-bond network.

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The Molten Globule, and Two-State vs. Non-Two-State Folding of Globular Proteins

Cited by 12 publications

References 92 publications

Probing Protein Folding with Sequence-Reversed α-Helical Bundles

Probing Protein Folding with Sequence-Reversed α-Helical Bundles

A conserved folding nucleus sculpts the free energy landscape of bacterial and archaeal orthologs from a divergent TIM barrel family

How coiled-coil assemblies accommodate multiple aromatic residues

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