LiteFold/DisProt huggingface dataset:
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 213
Annotation: negative regulation of cell population proliferation (Biological process)
Evidence: cell counting evidence used in manual assertion
Supporting statements:
The results of the colony-formation assay in soft agar medium demonstrated significantly greater colony formation in the control vector-transfected compared to the VHL‑expressing vector-transfected U87 GSLCs. 
Neurosphere formation, which is a reflection of the self‑renewal ability, was also significantly greater in the former than in the latter (P<(B) Results for neurosphere formation. After one week, soft agar colony formation of VHL gene‑transfected GSLCs was significantly
inhibited.0.001) 
 (A) Results on soft agar colony formation by control-vector and VHL-vector GSLC transfectants. Colony formation by the VHL transfectants was significantly inhibited. / (B) Results for neurosphere formation. After one week, soft agar colony formation of VHL gene‑transfected GSLCs was significantly inhibited.
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 213
Annotation: negative regulation of cell population proliferation (Biological process)
Evidence: cell proliferation assay evidence used in manual assertion
Supporting statements:
Proliferation of control vector-transfected U87 GSLCs was significantly more pronounced compared to the VHL gene-transfected GSLCs 7 days following transfection (P<0.01) (Fig. 4C), although the difference between them was smaller than in the case of the soft agar colony or neurosphere formation assay. 
(C) Results of cell-proliferation assay. After 7 days, control gene-transfected GSLCs showed significantly greater proliferation than the VHL gene-transfected GSLCs (P<0.01). * P<0.01, ** P<0.0
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 213
Annotation: negative regulation of signal transduction (Biological process)
Evidence: immunocytochemistry evidence used in manual assertion
Supporting statements:
These results suggest that VHL affected the JAK̸/STAT pathway as well as the PTEN/PI3K/Akt pathway; they also suggest that upregulation of PTEN by VHL gene transfer may affect the PI3K/Akt pathway, since PTEN is a PI3K/Akt pathway inhibitor
(A) Immunoreactivity rates for control-vector and VHL-vector glioma stem-like cell (GSLC) transfectants are shown. Expressions of CD133, STAT3,
JAK2 and Elongin A were significantly inhibited, whereas those of VHL and PTEN were significantly increased in VHL-transfectant GSLCs compared to
the control cells. Expressions of GFAP, NeuroD and MAP2 were not significantly altered. * P<0.05, ** P<0.01.
Protein: DP00287 (UniProt P40337)
Region: residues 97	to 102
Annotation: selective autophagy (Biological process)
Evidence: mutant phenotype evidence used in manual assertion
Supporting statements:
To determine whether the LIR motif of VHL specifically binds to LC3B, we generated point mutants of the VHL LIR motif (VHL-Y98H; VHL-L101A; VHL-Y98H and L101A, a double point mutant containing Y98H and L101A) using site-directed mutagenesis. Wild-type or mutant VHL and Flag-tagged LC3B were expressed, purified from Escherichia coli, and used in in vitro binding assay with the indicated proteins. Results show that the L101A mutation completely prevented the interaction between VHL and LC3B, whereas the histidine mutation at tyrosine 98 had no effect (Fig. 3g).
Taken together, the N-terminal LC3B and LIR motif of VHL interacted directly both in vitro and in vivo.
The annotated region includes the canonical LIR motif (YPTL) at positions 98-101.
Protein: DP00287 (UniProt P40337)
Region: residues 97	to 102
Annotation: protein binding (Molecular function)
Evidence: mutant phenotype evidence used in manual assertion
Supporting statements:
To determine whether the LIR motif of VHL specifically binds to LC3B, we generated point mutants of the VHL LIR motif (VHL-Y98H; VHL-L101A; VHL-Y98H and L101A, a double point mutant containing Y98H and L101A) using site-directed mutagenesis. Wild-type or mutant VHL and Flag-tagged LC3B were expressed, purified from Escherichia coli, and used in in vitro binding assay with the indicated proteins. Results show that the L101A mutation completely prevented the interaction between VHL and LC3B, whereas the histidine mutation at tyrosine 98 had no effect (Fig. 3g).
Taken together, the N-terminal LC3B and LIR motif of VHL interacted directly both in vitro and in vivo.
VHL directly binds to LC3B, the major marker of autophagy.
The annotated region includes the canonical LIR motif (YPTL) at positions 98-101.
Protein: DP00287 (UniProt P40337)
Region: residues 97	to 102
Annotation: protein binding (Molecular function)
Evidence: immunofluorescence confocal microscopy evidence used in manual assertion
Supporting statements:
Taken together, the N-terminal LC3B and LIR motif of VHL interacted directly both in vitro and in vivo.
VHL directly binds to LC3B, the major marker of autophagy.
We observed that LC3B co-localized with VHL in the cytosol (Fig. 3b). To determine the region of LC3B that binds to VHL, various truncations of LC3B were generated based on the sequence of the N-terminally Flag-tagged wild-type LC3B.
Protein: DP00287 (UniProt P40337)
Region: residues 97	to 102
Annotation: negative regulation of autophagy (Biological process)
Evidence: ubiquitination assay evidence used in manual assertion
Supporting statements:
In conclusion, our results showed that pVHL interacts with MAPL1LC3B and inhibits LC3B-mediated autophagy via MAP1LC3B ubiquitination. Furthermore, the activation of autophagy by the proteasome inhibitor MLN9708 induced cell death, indicating that MLN9708 can be used for VHL-deficient RCC therapy.
Polyubiquitination of LC3B is induced by the VHL E3 ubiquitin ligase.
We performed an in vitro ubiquitination assay using each indicated protein and an ATP-regeneration system. We observed that LC3B was polyubiquitinated by the VHL E3 ubiquitin complex in a time-dependent manner (Fig. 4b) and was subsequently degraded via the proteasome.
Protein: DP00287 (UniProt P40337)
Region: residues 97	to 102
Annotation: negative regulation of autophagy (Biological process)
Evidence: immunofluorescence confocal microscopy evidence used in manual assertion
Supporting statements:
In conclusion, our results showed that pVHL interacts with MAPL1LC3B and inhibits LC3B-mediated autophagy via MAP1LC3B ubiquitination. Furthermore, the activation of autophagy by the proteasome inhibitor MLN9708 induced cell death, indicating that MLN9708 can be used for VHL-deficient RCC therapy.
Polyubiquitination of LC3B is induced by the VHL E3 ubiquitin ligase.
Immunofluorescence staining was performed to identify the localization of LC3B and the 28S proteasome in cells with respect to VHL expression. Results show that Flag-LC3B and the 28S proteasome proteins co-localized more in VHL-deficient 786-o cells than in 786-o cells stably expressing VHL (Fig. 4f). This indicated that VHL ubiquitinated the N-terminal region (1−162 amino acids) of LC3B, and the leucine 101 in the LIR motif of VHL is essential for LC3B ubiquitination.
Protein: DP00287 (UniProt P40337)
Region: residues 54	to 157
Annotation: protein binding (Molecular function)
Evidence: yeast 2-hybrid evidence used in manual assertion
Supporting statements:
The pVHL β-domain is able to bind any member of the CDKN1 family, as indicated by yeast cell growth (Fig. 4B, Supplementary Figures S5 and S7). 
Yeast plasmids expressing either their N-terminal tail containing the CDI domain (p27-NT residues: 1–60; p21-NT: 1–49; p57-NT: 1–61) or the corresponding C-terminal moiety lacking the N-terminus (p27-ΔN residues: 61–198; p21-ΔN: 50–164; p57-ΔN: 62–316) were generated to test the effects of CDKN1 binding to pVHL30. As shown in Fig. 4D, loss of the p27 N-terminus clearly disrupts its ability to associate with pVHL30, as yeast cells expressing the C-terminus of p27 were all unable to grow in selective medium. Similar data have been also obtained for both p21 and p57 (Supplementary Figures S9 and S10), strongly supporting the notion that the CDI domain is responsible for CDKN1 binding to pVHL30.
Cross-references: DisProt:DP00016r041
DisProt:DP00018r056
DisProt:DP00017r018
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 213
Annotation: protein binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
Our data (Fig. 2B) indicates that all three CDKN1 proteins were able to interact with pVHL30, as demonstrated by their presence in the immunoprecipitate revealed with the anti-Flag antibody (bottom panels). Taken together, the experiments show that these proteins can form at least binary complexes in human, and notably kidney cells.
Cross-references: DisProt:DP00018r057
DisProt:DP00017r019
DisProt:DP00016r042
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 213
Annotation: DNA-binding transcription factor binding (Molecular function)
Evidence: yeast 2-hybrid evidence used in manual assertion
Supporting statements:
Expression in yeast cells of the Gal4-fusion proteins has been checked by Western blot analysis (Supplementary Figure S15), while the functional status of pVHL was verified testing the well-known pVHL/HIF-1α interaction with and without co-expressing PHD3 (Supplementary Figure S16).
Protein: DP00287 (UniProt P40337)
Region: residues 1	to 213
Annotation: protein serine/threonine kinase binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
As Aurora-A was co-immunoprecipitated with pVHL in all three cell lines (Figure 4C), we then asked whether pVHL was a phosphorylation substrate of Aurora-A.
Protein: DP00287 (UniProt P40337)
Region: residues 70	to 74
Annotation: phosphorylation display site (Disorder function)
Evidence: loss-of-function mutant phenotype evidence used in manual assertion
Supporting statements:
This annotation refers to the phosphorylation of the Ser 72 residue of VHL protein by human Aurora kinase A.
Mutation of Ser72 into alanine (VHL SA72) greatly diminished phosphorylation of pVHL by Aurora-A (Figure 5, lane 2).
Protein: DP00288 (UniProt Q06253)
Region: residues 52	to 73
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
Far UV CD experiments show that Phd52–73Se is intrinsically unstructured in its isolated state but gains an appreciable amount of α-helix upon binding to Doc (Fig. 3).
Protein: DP00288 (UniProt Q06253)
Region: residues 52	to 73
Annotation: disorder to order (Structural transition)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
Far UV CD experiments show that Phd52–73Se is intrinsically unstructured in its isolated state but gains an appreciable amount of α-helix upon binding to Doc (Fig. 3).
Protein: DP00288 (UniProt Q06253)
Region: residues 52	to 73
Annotation: protein binding (Molecular function)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
The action of Doc is suppressed by the antitoxin Phd, which consists of two domains. Its C-terminal domain (residues 52–73) harbors the interaction site with Doc and on its own prevents Doc-mediated growth arrest
Far UV CD experiments show that Phd52–73Se is intrinsically unstructured in its isolated state but gains an appreciable amount of α-helix upon binding to Doc (Fig. 3).
Protein: DP00288 (UniProt Q06253)
Region: residues 41	to 73
Annotation: disorder (Structural state)
Evidence: small-angle X-ray scattering evidence used in manual assertion
Supporting statements:
The plateau at I(s)s2 > 0 observed in the Kratky plot (Figure 1D) indicates that Phd is partially unstructured (Pollack et al., 1999; Porod, 1982), in agreement with earlier circular dichroism (CD) studies (Gazit and Sauer, 1999).
 Using the ensemble optimization method (EOM) (Bernado et al., 2007), we find that the scattering curve can be modeled assuming a fixed conformation for residues 1–40 of Phd and a wide structural ensemble for residues 41–73 (c2 = 0.8, Figure 1D, Table S1).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
The amount of secondary structure was estimated from the CD spectrum (20, 21). At 37 °C in buffer alone, Phd seems to be in a largely unfolded, random-coil conformation. However, at 4 °C or at 37 °C in the presence of trimethylamine N-oxide, Phd has a CD spectra indicative of a folded protein containing approximately 45% a-helix (Fig. 1A).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: disorder to order (Structural transition)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
DNA binding induces a dose-dependent increase in Phd structure, suggesting that the folding and site-specific DNA binding of Phd are coupled.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: protein binding (Molecular function)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
The slope of the initial part of the titration curve shows that the amount of operator DNA needed for induction of fully folded Phd is roughly one-quarter of the protein concentration, suggesting that Phd binds to its operator DNA as a tetramer.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: DNA-binding transcription repressor activity (Molecular function)
Evidence: electrophoretic mobility shift assay evidence used in manual assertion
Supporting statements:
Fig. 5,A and B, show a gel-mobility-shift assay of Phd binding to DNA fragments containing the left and the right operator subsites, respectively. The assays were performed using a DNA concentration low enough (10 pM) to allow the reaction mechanism to be deduced from the dependence of binding on Phd concentration and an equilibrium dissociation constant to be calculated. Two observations suggest that two Phd monomers bind cooperatively to subsite DNA.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: DNA-binding transcription repressor activity (Molecular function)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
DNA binding induces a dose-dependent increase in Phd structure, suggesting that the folding and site-specific DNA binding of Phd are coupled.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 57
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy-based hydrogen-deuterium exchange evidence used in manual assertion
Supporting statements:
When attempting to probe the flexibility of Phd1–57 with hydrogen-deuterium exchange experiments, virtually all amide protons exchange within the set-up time (∼11 min) prior to the start of NMR acquisition, and their 1H-15N cross-peaks are therefore absent in the HSQC spectrum. Such dramatic behavior indicates very low stability and lack of conformational rigidity and is presumably the result of a high overall conformational exchange of Phd between folded and partially folded states (See Extended Experimental Procedures for further details).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 57
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
We used NMR to characterize the structure of the N-terminal domain of Phd (Phd1–57—residues 1–57) in solution. This N-terminal domain of Phd examined by two-dimensional (2D) 1H-15N HSQC NMR is in agreement with a conformationally heterogeneous protein that populates folded and disordered states (Figure S1F, in cyan).
Given the expected elevated conformational heterogeneity of Phd1–57, the 15N transverse and longitudinal relaxation rates (R2 and R1) are the best indicators to probe the state of the Phd1–57 ensemble in solution. Figure 1E shows the R2/R1 ratios along the Phd1–57 sequence (represented as red bars). Two regions show significantly high R2/R1 ratios as a result of elevated R2 rates, indicative of a pronounced conformational freedom and high exchange between different conformational states in the μs to ms timescale. Additionally the C-terminal region displays pronounced low R2/R1 ratios, which indicate an increase in fast dynamics (i.e., high degree of flexibility) in agreement with the heteronuclear NOE values found for this region of the domain (Figure S1G).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: disorder (Structural state)
Evidence: differential scanning calorimetry evidence used in manual assertion
Supporting statements:
In its unbound state Phd is only marginally stable (Figure 2B). 
This indicates a poorly structured unfolding unit, suggesting that, in agreement with the NMR data, the partly unfolded species dominates in solution.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 57
Annotation: disorder to order (Structural transition)
Evidence: differential scanning calorimetry evidence used in manual assertion
Supporting statements:
Differential scanning calorimetry (DSC) unfolding data of Phd and its N-terminal fragment Phd1–57 fit well to a two-state model consisting of a dimer unfolding into monomer species. In its unbound state Phd is only marginally stable (Figure 2B).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 57
Annotation: protein binding (Molecular function)
Evidence: differential scanning calorimetry evidence used in manual assertion
Supporting statements:
Differential scanning calorimetry (DSC) unfolding data of Phd and its N-terminal fragment Phd1–57 fit well to a two-state model consisting of a dimer unfolding into monomer species. In its unbound state Phd is only marginally stable (Figure 2B).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: protein binding (Molecular function)
Evidence: differential scanning calorimetry evidence used in manual assertion
Supporting statements:
Our DSC measurements show that binding of DNA or of its toxin partner Doc to Phd significantly increases its thermodynamic stability (Figure 2B, Figure S2C, and Table S2). In both cases the increment in the conformational stability is accompanied by a large increase in the heat of unfolding and the melting temperature of Phd.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: DNA binding (Molecular function)
Evidence: differential scanning calorimetry evidence used in manual assertion
Supporting statements:
Our DSC measurements show that binding of DNA or of its toxin partner Doc to Phd significantly increases its thermodynamic stability (Figure 2B, Figure S2C, and Table S2). In both cases the increment in the conformational stability is accompanied by a large increase in the heat of unfolding and the melting temperature of Phd.
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: protein binding (Molecular function)
Evidence: isothermal titration calorimetry evidence used in manual assertion
Supporting statements:
The effect of Doc on the affinity of Phd for a single DNA-binding site (OR1) was quantified using isothermal titration calorimetry (ITC). The affinity of Phd2Doc for OR1 is 0.3 μM, about 10-fold higher than the binding constant of Phd2 for OR1 (Figures 3D and 3E; Table S4).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: DNA binding (Molecular function)
Evidence: isothermal titration calorimetry evidence used in manual assertion
Supporting statements:
The effect of Doc on the affinity of Phd for a single DNA-binding site (OR1) was quantified using isothermal titration calorimetry (ITC). The affinity of Phd2Doc for OR1 is 0.3 μM, about 10-fold higher than the binding constant of Phd2 for OR1 (Figures 3D and 3E; Table S4).
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 58
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Phd crystallizes in two different crystal forms. Form I contains a version of the protein that is truncated at Ser58 (hereafter termed Phd1–57). All four molecules in the asymmetric unit adopt a similar, well-folded conformation encompassing Met1-Phe56. This N-terminal domain forms a small globular homodimer comprising a central six-stranded β sheet decorated with four α helices (two from each monomer), resembling the antitoxin YefM (Kamada and Hanaoka, 2005
) (Figure 1A ; Figure S1A available online). The N-terminal α helices α1 and α2 form a large positively charged surface that constitutes the DNA-binding region (Figure S1B). The dimer interface is relatively small (about 900 Å2 of water accessible surface buried per dimer).
Cross-references: PDB:3HS2
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 58
Annotation: disorder to order (Structural transition)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Phd crystallizes in two different crystal forms. Form I contains a version of the protein that is truncated at Ser58 (hereafter termed Phd1–57). All four molecules in the asymmetric unit adopt a similar, well-folded conformation encompassing Met1-Phe56. This N-terminal domain forms a small globular homodimer comprising a central six-stranded β sheet decorated with four α helices (two from each monomer), resembling the antitoxin YefM (Kamada and Hanaoka, 2005) (Figure 1A ; Figure S1A available online). The N-terminal α helices α1 and α2 form a large positively charged surface that constitutes the DNA-binding region (Figure S1B). The dimer interface is relatively small (about 900 Å2 of water accessible surface buried per dimer).
In this case the disordered nature of the quoted region does not (only) come from missing residues but also from the large differences between the coordinates of the various copies of the protein in the crystal. This shows that some residues have resolved coordinates only because of crystal contacts, and these have been annotated as disordered as well (in line with the authors' assertion).
Cross-references: PDB:3HS2
Protein: DP00288 (UniProt Q06253)
Region: residues 56	to 73
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
About 25% of the surface of Doc is buried upon interacting with two Phd dimers (Figure 4B). Site H corresponds to the one identified earlier in the crystal structure of a nontoxic mutant of Doc (Garcia-Pino et al., 2008) and recognizes the Phd segment Phe56-Arg73.
Cross-references: PDB:3K33
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: disorder to order (Structural transition)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
The asymmetric unit of our Phd/Doc cocrystals contains three copies of Phd and one copy of Doc, which through crystal symmetry form a hetero-octameric Phd2-Doc-Phd2-Doc-Phd2 complex (Figure 4A ; Figure S4A).
While the authors do not explicitly state this, the structured state of phd becomes evident when looking at the corresponding complex structure in PDB:3k33.
Cross-references: PDB:3K33
Protein: DP00288 (UniProt Q06253)
Region: residues 1	to 73
Annotation: protein binding (Molecular function)
Evidence: surface plasmon resonance evidence used in manual assertion
Supporting statements:
Using surface plasmon resonance (SPR), we measured the binding kinetics and affinities for both interaction modes between Phd and Doc (Figure 2C). Consistent with the crystal structure of the complex, the best model that explains the binding data involves two Phd dimers binding to a Doc monomer with affinities of 0.35 μM and 31 μM (Figure 2C; Table S4).