LiteFold/DisProt huggingface dataset:
Protein: DP00285 (UniProt P10388)
Region: residues 168	to 461
Annotation: pre-molten globule (Structural state)
Evidence: dynamic light scattering assay evidence used in manual assertion
Supporting statements:
The ROA spectrum of the T-A-1 peptide indicates the presence of a large amount of PPII helix, which, although it might be expected to promote an extended structure, is apparently imperfect to some degree, as indicated by a hydrodynamic radius of 4.0 nm as measured by dynamic light scattering under the same conditions of pH and temperature as for the ROA spectra. This corresponds to the hydrodynamic radius expected for a natively unfolded premolten-globular protein of the same molecular weight ( 44).
Protein: DP00286 (UniProt Q32NN2)
Region: residues 203	to 215
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
By contrast, the relaxation parameters for the C-terminal QUA2 region indicate substantial backbone flexibility. In particular, the decreased 15N R2 values and the large negative NOEs observed for residues 183–188 and 202–214 (Figure 3) are characteristic of extensive local motion on the sub-nanosecond timescale.
Cross-references: PDB:2BL5
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 59
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
VHL residues 1–59 are disordered and not observed in the structure.
Cross-references: PDB:4WQO
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: molten globule (Structural state)
Evidence: near-UV circular dichroism evidence used in manual assertion
Supporting statements:
Yet, the near-UV CD experiments show an absence of a tertiary structure.
We therefore conclude that, under native conditions, the non-bound pVHL has a molten globule configuration with marginal stability.
The near-UV CD spectra of pVHL at 25 °C shown in Fig. 1B reveals no significant signals originating from aromatic side chains, thus suggesting a dramatic loss of tertiary structure of the soluble and unbound protein in comparison with the complex-related crystal structure.
Here, we present clear evidence of a molten globule conformation of the pVHL tumor suppressor protein under physiologically relevant solvent conditions.
The indications of a molten globule structure of pVHL are clear, given that the protein possesses a typical combination of an apparent secondary structure (Fig. 2A) with no tertiary structure
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: molten globule (Structural state)
Evidence: pH-induced protein unfolding evidence used in manual assertion
Supporting statements:
Upon the addition of urea, even at very low concentrations, the protein unfolds in a non-reversible manner, leading to the formation of amorphous aggregates.
We therefore conclude that, under native conditions, the non-bound pVHL has a molten globule configuration with marginal stability.
The degree of denaturation was assessed by monitoring ellipticity changes at 222 and 218 nm as a function of urea concentrations. As can be observed in Fig. 2, the unfolding of pVHL occurs at fairly low urea concentrations. Unfolding as reflected by a significant reduction in ellipticity was observed at urea concentrations as low as 0.25 m at both temperatures. Furthermore, the unfolding event was partially irreversible, and the formation of amorphous aggregates could be visualized by the naked eye.
However, the linear nature of the initial slope of the curve and the aggregative behavior implies that unbound pVHL stability is marginal, as was observed previously with other tumor suppressor proteins
The thermodynamic stability of the secondary structure as observed by CD is also marginal; at very low concentrations of urea the protein undergoes denaturation (Fig. 2), which results in some extent of aggregation.
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: molten globule (Structural state)
Evidence: chromatography evidence used in manual assertion
Supporting statements:
Dynamic light-scattering and gel filtration chromatography of the purified pVHL clearly indicated that the Stokes radius of the protein is larger than what would be expected from its crystal structure. However, under these conditions, the protein shows a clear secondary structure as determined by far-UV circular dichroism.
We therefore conclude that, under native conditions, the non-bound pVHL has a molten globule configuration with marginal stability.
Size exclusion chromatography was therefore used to estimate the hydrodynamic dimensions of the protein and as a probe for elucidating the compactness of the protein's tertiary structure, because the elution volume of partially or fully unfolded proteins is significantly smaller than that of well folded proteins due to the large increase in the Stokes radius.
Fig. 3A shows the Kav versus Mw plot; the calculated Kav for pVHL was 0.191, corresponding to a molecular mass of 47,130 Da (calculated with curve's equation), which is larger than the theoretical molecular mass value of pVHL (19,000 Da). Fig. 3B shows √-logKav versus Stokes radius (RS); according to this relation, the calculated RS for pVHL is 30.8 Å.
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: molten globule (Structural state)
Evidence: dynamic light scattering assay evidence used in manual assertion
Supporting statements:
Dynamic light-scattering and gel filtration chromatography of the purified pVHL clearly indicated that the Stokes radius of the protein is larger than what would be expected from its crystal structure. However, under these conditions, the protein shows a clear secondary structure as determined by far-UV circular dichroism.
We therefore conclude that, under native conditions, the non-bound pVHL has a molten globule configuration with marginal stability.
The measured hydrodynamic radius was 3 nm, and the estimated molecular mass was 43 kDa. These values, taken together with the size exclusion chromatography results, are significantly larger for the solution form of the protein than the values obtained from the folded pVHL crystal structure. This could indicate a somewhat flexible and non-compact conformation for the unbound protein.
The percent of scattered mass as attributed to the apparent hydrodynamic radius. RH for pVHL is 3 nm and estimated Mw is 43 kDa.
Here, we present clear evidence of a molten globule conformation of the pVHL tumor suppressor protein under physiologically relevant solvent conditions.
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: molten globule (Structural state)
Evidence: fluorescence evidence used in manual assertion
Supporting statements:
The fluorescence of the hydrophobic ANSA probe is much stronger in the presence of molten globular state of proteins than with a rigid, well folded, or fully unfolded state (36). Fig. 5 shows that, upon binding to pVHL, ANSA fluorescence intensity increases by 50-fold, whereas the emission spectrum indicates a blue shift toward 400 nm. This could suggest that the hydrophobic core exists, but in a rather loosely packed conformation, and is therefore consistent with a molten globule tertiary structure.
Furthermore, such a significant increase in ANSA fluorescence (Fig. 5) clearly indicates a molten globule structure rather than the existence of a monomeric or dimeric well folded protein or, alternatively, an unfolded one.
Here, we present clear evidence of a molten globule conformation of the pVHL tumor suppressor protein under physiologically relevant solvent conditions.
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: molten globule (Structural state)
Evidence: analytical ultracentrifugation evidence used in manual assertion
Supporting statements:
The sedimentation equilibrium data fit well to a single species with a calculated apparent molecular mass of 18,200 Da, implying that pVHL is monomeric in solution. This agrees very well with the Mr established by SDS gel electrophoresis and the Mr of 8,500 Da predicted for pVHL from the DNA sequence. There was some degree of aggregation, as observed at the lower part of the tube. This may reflect a moderate degree of nonspecific aggregation as may be expected for a partially unfolded protein after long incubation at high concentration.
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 213
Annotation: intracellular non-membrane-bounded organelle (Cellular component)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
Upon the addition of urea, even at very low concentrations, the protein unfolds in a non-reversible manner, leading to the formation of amorphous aggregates.
The degree of denaturation was assessed by monitoring ellipticity changes at 222 and 218 nm as a function of urea concentrations. As can be observed in Fig. 2, the unfolding of pVHL occurs at fairly low urea concentrations. Unfolding as reflected by a significant reduction in ellipticity was observed at urea concentrations as low as 0.25 m at both temperatures. Furthermore, the unfolding event was partially irreversible, and the formation of amorphous aggregates could be visualized by the naked eye.
However, the linear nature of the initial slope of the curve and the aggregative behavior implies that unbound pVHL stability is marginal, as was observed previously with other tumor suppressor proteins
The thermodynamic stability of the secondary structure as observed by CD is also marginal; at very low concentrations of urea the protein undergoes denaturation (Fig. 2), which results in some extent of aggregation.
Protein: DP00287 (UniProt P40337)
Region: residues 67	to 117
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
pVHL binds to HIF only when a conserved proline in HIF is hydroxylated, a modification that is oxygen-dependent. The 1.85 angstrom structure of a 20-residue HIF-1α peptide–pVHL–ElonginB–ElonginC complex shows that HIF-1α binds to pVHL in an extended β strand–like conformation. The hydroxyproline inserts into a gap in the pVHL hydrophobic core, at a site that is a hotspot for tumorigenic mutations, with its 4-hydroxyl group recognized by buried serine and histidine residues.
To investigate the targeting of HIF-1α by the VBC-CR ubiquitin-protein ligase and the basis of hydroxyproline recognition in intracellular signaling, we determined the 1.85 Å crystal structure of the pVHL–ElonginB–ElonginC (VBC) complex bound to the hydroxyproline-containing 20-residue destruction sequence of HIF-1α (Table 1 and fig. S1). The structure shows that a 15–amino acid portion of HIF-1α (residues 561 to 575) adopts an extended, β strand–like conformation (Fig. 1A). It binds pVHL in a bipartite manner, with two discontinuous HIF-1α segments interacting with a continuous site on pVHL.
HIF-1α interacts exclusively with the β domain of pVHL. It binds alongside the β sandwich, making five backbone-backbone hydrogen bonds (Fig. 1A). The side of the pVHL β sandwich where HIF-1α binds has the hydrophobic core partially exposed. This exposed hydrophobic patch, together with several partially buried polar residues, makes up the binding site of the hydroxyproline.
The hydroxyproline has a central role in complex formation. It is nearly entirely buried, with 96% of its accessible surface area in a hypothetical free peptide covered by pVHL. The pyrrolidine ring inserts toward the partially exposed hydrophobic core of the pVHL β domain, making multiple van der Waals contacts with Trp88, Tyr98, and Trp117 of pVHL (Fig. 2A). The 4-hydroxyl group inserts farthest into pVHL and forms hydrogen bonds with the Nδ of His115 (2.7 Å) and the OH group of Ser111 (2.7 Å), both of which also form hydrogen bonds with other pVHL groups (Fig. 2A). His115 and Ser111 are partially solvent exposed in the apo-VBC structure (10) but become entirely buried on HIF-1α binding.
The dotted lines indicate hydrogen bonds between the Gln67Oδ1, Tyr98 Oη, His110 NH, and His110 CO groups of pVHL, and the Leu562 NH, Hyp564 CO, Tyr565 NH, and Tyr565 CO groups of HIF-1α.
Compared with Hyp564, the other N-segment residues make significantly fewer contacts. Among them, Ile566 makes the most contacts, interacting with Pro99 and Ile109 of pVHL. Met561 packs with Phe91 of pVHL, Leu562 with Tyr112and Arg69, Ala563 with Trp88, and Tyr565 with His110 (Fig. 2A).
pVHL residues contacting HIF1α: N67, R69, W88, F91, Y98, P99, I109, H110, S111, Y112, H115, W117.
Cross-references: PDB:1LM8
Protein: DP00287 (UniProt P40337)
Region: residues 156	to 161
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
To investigate the targeting of HIF-1α by the VBC-CR ubiquitin-protein ligase and the basis of hydroxyproline recognition in intracellular signaling, we determined the 1.85 Å crystal structure of the pVHL–ElonginB–ElonginC (VBC) complex bound to the hydroxyproline-containing 20-residue destruction sequence of HIF-1α (Table 1 and fig. S1).
The α domain binds ElonginC, which, in association with ElonginB, recruits the VBC complex to Cul2-Rbx1 to form the ubiquitin-protein ligase (8–10).
pVHL residues Leu158 and Lys159 contact residue Cys112 of Elongin C.
Protein: DP00287 (UniProt P40337)
Region: residues 80	to 84
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Mutation of the VHL tumor suppressor is associated with the inherited von Hippel–Lindau (VHL) cancer syndrome and the majority of kidney cancers. VHL binds the ElonginC-ElonginB complex and regulates levels of hypoxia-inducible proteins. The structure of the ternary complex at 2.7 angstrom resolution shows two interfaces, one between VHL and ElonginC and another between ElonginC and ElonginB. Tumorigenic mutations frequently occur in a 35-residue domain of VHL responsible for ElonginC binding. A mutational patch on a separate domain of VHL indicates a second macromolecular binding site.
The VHL-ElonginC interface is almost completely hydrophobic with only a handful of significant hydrogen bonds at the periphery (Fig. 3A). The H1 helix of the VHL α domain fits into the concave surface of ElonginC, inserting hydrophobic side chains into pockets along the surface (Fig. 3A). Reciprocally, the H4 helix of ElonginC, which bulges out from the side of the concave surface, fits into an extended groove formed by the H1, H2, and H3 helices of the VHL α domain and completes the intermolecular four-helix cluster packing (Fig. 3A).
The H1 helix of the VHL α domain coincides with the 12–amino acid segment shown to be important for ElonginC binding (10), and the structure reveals that it makes extensive contacts to ElonginC. The most significant van der Waals contacts are made by Leu158, which protrudes from the H1 helix and fits into an ElonginC pocket, and by Cys162 and Arg161 (Fig. 3A). These are augmented by contacts from the Lys159, Val165, Val166, and Leu169 side chains of VHL (Fig. 3A). The other two helices of the α domain also contribute contacts (Leu178, Ile180, and Leu184), with Leu184 making the most extensive ones in this region (Fig. 3A). Additional contacts are made by residues in the first α-β linker (Leu153 and Val155) and by Arg82 from the β domain (Fig. 3A). The hydrogen bonds made by Arg82, together with those made by Lys159 and Arg161 from the H1 helix, represent the few significant hydrogen-bond contacts made at the VHL-ElonginC interface. The arginine side chains are also anchored in the hydrogen-bond networks of the VHL α-β domain interface (Fig. 2D). 
pVHL residues contacting Elongin C: R82, L153, V155, Y156, L158, K159, R161, V165, V166, L169, L178, I180, L184
This evidence corresponds to the Arg82 site from pVHL β domain.
Cross-references: PDB:1VCB
Protein: DP00287 (UniProt P40337)
Region: residues 153	to 184
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Mutation of the VHL tumor suppressor is associated with the inherited von Hippel–Lindau (VHL) cancer syndrome and the majority of kidney cancers. VHL binds the ElonginC-ElonginB complex and regulates levels of hypoxia-inducible proteins. The structure of the ternary complex at 2.7 angstrom resolution shows two interfaces, one between VHL and ElonginC and another between ElonginC and ElonginB. Tumorigenic mutations frequently occur in a 35-residue domain of VHL responsible for ElonginC binding. A mutational patch on a separate domain of VHL indicates a second macromolecular binding site.
The VHL-ElonginC interface is almost completely hydrophobic with only a handful of significant hydrogen bonds at the periphery (Fig. 3A). The H1 helix of the VHL α domain fits into the concave surface of ElonginC, inserting hydrophobic side chains into pockets along the surface (Fig. 3A). Reciprocally, the H4 helix of ElonginC, which bulges out from the side of the concave surface, fits into an extended groove formed by the H1, H2, and H3 helices of the VHL α domain and completes the intermolecular four-helix cluster packing (Fig. 3A).
The H1 helix of the VHL α domain coincides with the 12–amino acid segment shown to be important for ElonginC binding (10), and the structure reveals that it makes extensive contacts to ElonginC. The most significant van der Waals contacts are made by Leu158, which protrudes from the H1 helix and fits into an ElonginC pocket, and by Cys162 and Arg161 (Fig. 3A). These are augmented by contacts from the Lys159, Val165, Val166, and Leu169 side chains of VHL (Fig. 3A). The other two helices of the α domain also contribute contacts (Leu178, Ile180, and Leu184), with Leu184 making the most extensive ones in this region (Fig. 3A). Additional contacts are made by residues in the first α-β linker (Leu153 and Val155) and by Arg82 from the β domain (Fig. 3A). The hydrogen bonds made by Arg82, together with those made by Lys159 and Arg161 from the H1 helix, represent the few significant hydrogen-bond contacts made at the VHL-ElonginC interface. The arginine side chains are also anchored in the hydrogen-bond networks of the VHL α-β domain interface (Fig. 2D). 
pVHL residues contacting Elongin C: R82, L153, V155, Y156, L158, K159, R161, V165, V166, L169, L178, I180, L184
Cross-references: PDB:1VCB
Protein: DP00287 (UniProt P40337)
Region: residues 153	to 184
Annotation: regulation of cellular response to hypoxia (Biological process)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Mutation of the VHL tumor suppressor is associated with the inherited von Hippel–Lindau (VHL) cancer syndrome and the majority of kidney cancers. VHL binds the ElonginC-ElonginB complex and regulates levels of hypoxia-inducible proteins. The structure of the ternary complex at 2.7 angstrom resolution shows two interfaces, one between VHL and ElonginC and another between ElonginC and ElonginB. Tumorigenic mutations frequently occur in a 35-residue domain of VHL responsible for ElonginC binding. A mutational patch on a separate domain of VHL indicates a second macromolecular binding site.
Protein: DP00287 (UniProt P40337)
Region: residues 153	to 184
Annotation: molecular adaptor activity (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Mutation of the VHL tumor suppressor is associated with the inherited von Hippel–Lindau (VHL) cancer syndrome and the majority of kidney cancers. VHL binds the ElonginC-ElonginB complex and regulates levels of hypoxia-inducible proteins. The structure of the ternary complex at 2.7 angstrom resolution shows two interfaces, one between VHL and ElonginC and another between ElonginC and ElonginB. Tumorigenic mutations frequently occur in a 35-residue domain of VHL responsible for ElonginC binding. A mutational patch on a separate domain of VHL indicates a second macromolecular binding site.
Protein: DP00287 (UniProt P40337)
Region: residues 67	to 117
Annotation: molecular adaptor activity (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
To investigate the targeting of HIF-1α by the VBC-CR ubiquitin-protein ligase and the basis of hydroxyproline recognition in intracellular signaling, we determined the 1.85 Å crystal structure of the pVHL–ElonginB–ElonginC (VBC) complex bound to the hydroxyproline-containing 20-residue destruction sequence of HIF-1α (Table 1 and fig. S1). The structure shows that a 15–amino acid portion of HIF-1α (residues 561 to 575) adopts an extended, β strand–like conformation (Fig. 1A). It binds pVHL in a bipartite manner, with two discontinuous HIF-1α segments interacting with a continuous site on pVHL.
Protein: DP00287 (UniProt P40337)
Region: residues 72	to 78
Annotation: amyloid fibril formation (Biological process)
Evidence: transmission electron microscopy evidence used in manual assertion
Supporting statements:
Here we report that using in silico approaches, we identified six (3-9 residues long) fragments predicted to harbor propensity to form amyloid fibrils. By a series of complementary biophysical techniques, including ThT binding assay, CD spectroscopy, Congo Red birefringence, transmission electron microscopy (TEM), atomic force microscopy (AFM), and X-ray diffraction, we found that two of the fragments, 72SQVIFCN78 and 147IFANITL153 , self-assembled in vitro and formed canonical amyloid fibrils β-sheets rich structures.
We performed TEM analysis to visualize the morphology of the SN7 and IL7 assemblies. As shown in Figure 2b and 2c, the assemblies exhibited fibrillar morphology for SN7 and IL7 aggregates, respectively.
Protein: DP00287 (UniProt P40337)
Region: residues 147	to 153
Annotation: amyloid fibril formation (Biological process)
Evidence: transmission electron microscopy evidence used in manual assertion
Supporting statements:
Here we report that using in silico approaches, we identified six (3-9 residues long) fragments predicted to harbor propensity to form amyloid fibrils. By a series of complementary biophysical techniques, including ThT binding assay, CD spectroscopy, Congo Red birefringence, transmission electron microscopy (TEM), atomic force microscopy (AFM), and X-ray diffraction, we found that two of the fragments, 72SQVIFCN78 and 147IFANITL153 , self-assembled in vitro and formed canonical amyloid fibrils β-sheets rich structures.
We performed TEM analysis to visualize the morphology of the SN7 and IL7 assemblies. As shown in Figure 2b and 2c, the assemblies exhibited fibrillar morphology for SN7 and IL7 aggregates, respectively.
Protein: DP00287 (UniProt P40337)
Region: residues 147	to 153
Annotation: amyloid fibril formation (Biological process)
Evidence: atomic force microscopy evidence used in manual assertion
Supporting statements:
Here we report that using in silico approaches, we identified six (3-9 residues long) fragments predicted to harbor propensity to form amyloid fibrils. By a series of complementary biophysical techniques, including ThT binding assay, CD spectroscopy, Congo Red birefringence, transmission electron microscopy (TEM), atomic force microscopy (AFM), and X-ray diffraction, we found that two of the fragments, 72SQVIFCN78 and 147IFANITL153 , self-assembled in vitro and formed canonical amyloid fibrils β-sheets rich structures.
Furthermore, we performed AFM imaging to get additional details regarding the structural parameters such as height and diameter of the amyloid fibrils. Representative AFM images of amyloids formed by SN7 and IL7 are shown in Figure 2e, 2f, respectively. The amyloids of SN7 appeared as clusters of intercalated fibrils with a height of 5-10 nm and of diameter ~30 nm (n=30). The IL7 amyloids were observed to be of 3-13 nm height with a diameter of ~46 nm and varying length (n=30).