LiteFold/DisProt huggingface dataset:
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: protein binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
We showed that RNF126 interacts with p21 and RNF126 overexpression increased p21
protein ubiquitination in an E3 ligase activity-dependent manner. 
When
Flag-RNF126 was immunoprecipitated by anti-Flag Ab, the
endogenous p21 protein could be detected in the protein
complex (Fig. 3A). In addition, the protein–protein interaction
between endogenous RNF126 and endogenous p21 was
detected in MDA-MB-231 (Fig. 3B). These results suggest that
RNF126 and p21 proteins interact with each other under
physiologic conditions. 
RNF126 and p21 proteins interact with each other.
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: ubiquitination display site (Disorder function)
Evidence: immunoprecipitation evidence used in manual assertion
Supporting statements:
These ﬁndings suggest that RNF126 promotes cancer cell proliferation by targeting p21 for ubiquitin-mediated degradation.
Protein: DP00016 (UniProt P38936)
Region: residues 139	to 160
Annotation: protein binding (Molecular function)
Evidence: enzyme-linked immunoabsorbent assay evidence used in manual assertion
Supporting statements:
To demonstrate p21Cip1 inhibition of the binding of the C-terminal domain of p66 to PCNA, the wells were coated with a streptavidin-biotinated p66 peptide complex by adding 50 μl of 20 μg/ml streptavidin plus 100 ng of biotinated p66 peptide in PBS to each well for 1 h.
Protein: DP00016 (UniProt P38936)
Region: residues 139	to 160
Annotation: protein sequestering activity (Molecular function)
Evidence: protein detection assay evidence used in manual assertion
Supporting statements:
Conversely, p66 interacts with the domain-interconnecting loop of PCNA, a region previously shown to be important for DNA polymerase delta activity and for binding of the cell cycle inhibitor p21 Cip1 . In accordance with this, a peptide containing the PCNA-binding domain of p21 Cip1 inhibited p66 binding to PCNA and the activity of native three-subunit DNA polymerase delta
In Fig. 4, we show that a peptide of 22 amino acids containing the PCNA-binding domain of p21 Cip1 inhibits both p66 peptide binding to PCNA and DNA synthesis by a mouse DNA polymerase delta fraction containing p66 (30) (Fig. 4B, inset).
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: negative regulation of cell population proliferation (Biological process)
Evidence: cell growth regulation assay evidence used in manual assertion
Supporting statements:
Cell death was greatest in MT1A2 and A549 cells transduced with Ad vectors expressing p16, p18 and p27 (Figure 2 and Table 2), whereas p19 and p21 expression had a more cytostatic effect on cell proliferation.
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: molecular function activator activity (Molecular function)
Evidence: small interfering RNA knockdown evidence used in manual assertion
Supporting statements:
Additionally, we demonstrate that fibrillar collagen can also arrest cells at the G 2 phase, which is mediated in part by p21 CIP1
p21 CIP1 siRNA alone had a minimal effect on cell cycle distribution and did not significantly increase cell proliferation (data not shown), indicating that it is the cooperation between p15 INK4b and p21 CIP1 that is responsible for the cell cycle arrest observed when M24met cells are plated in the presence of FC. 
FC means fibrillar collagen
Protein: DP00016 (UniProt P38936)
Region: residues 152	to 158
Annotation: protein binding (Molecular function)
Evidence: glutathione S-transferase pull-down assay evidence used in manual assertion
Supporting statements:
Data from GST pull-down experiments suggest that the C-terminal region (amino acids 152–158) of p21 mediates TRIM39-p21 association (Fig. 2C).
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: protein binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
Immunoprecipitation and immunoblotting confirmed that cyclin D1 and p21CIP1 were associated with CDK4 and CDK6 in complexes of this size (see Fig. S1 in the supplemental material; also data not shown).
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: protein binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
Mechanistically, TRIM39 interacts with p21, which subsequently prevents Cdt2 from binding to p21, therefore blocking ubiquitylation and proteasomal degradation of p21 mediated by CRL4(Cdt2) E3 ligase. 
To test if TRIM39 associates with endogenous p21 protein, we introduced TRIM39α-Myc or TRIM39β-Myc into HCT116 WT cells and then performed immunoprecipitation with anti-Myc antibody. The association between p21 and TRIM39α or TRIM39β was readily detected (Fig. 2B)
In this study, we identiﬁed TRIM39 as a binding partner of p21 in the nucleus. Exogenous TRIM39 stabilized p21 by preventing its ubiquitylation, whereas TRIM39 depletion resulted in destabilization of p21, which was accompanied by increased ubiquitylation.
Protein: DP00016 (UniProt P38936)
Region: residues 143	to 147
Annotation: phosphorylation display site (Disorder function)
Evidence: loss-of-function mutant phenotype evidence used in manual assertion
Supporting statements:
Inactivation of Thr 145 markedly reduced the Akt-dependent phosphorylation of p21Cip1 (Fig.2A). This suggests that Thr 145 serves as the Akt phosphorylation acceptor amino acid.
This annotation refers to the phosphorylation of the Thr 145 residue, included in the consensus motif for Akt-mediated phosphorylation MOD_PKB_1 that comprises the residues 140 to 148.
Cross-references: ELM:MOD_PKB_1
Protein: DP00016 (UniProt P38936)
Region: residues 143	to 147
Annotation: phosphorylation display site (Disorder function)
Evidence: in vitro protein kinase assay evidence used in manual assertion
Supporting statements:
In vitro kinase assay analysis confirms that Akt and PKA both phosphorylate p21Cip1 at the same site (Thr 145) (Fig. ​2B). 
This annotation refers to the phosphorylation of the Thr 145 residue, included in the consensus motif for Akt-mediated phosphorylation MOD_PKB_1 that comprises the residues 140 to 148.
Cross-references: ELM:MOD_PKB_1
Protein: DP00016 (UniProt P38936)
Region: residues 139	to 160
Annotation: protein binding (Molecular function)
Evidence: X-ray crystallography-based structural model with high relative B-factor values used in manual assertion
Supporting statements:
A 22 residue peptide is bound to each of the three PCNA subunits of the ring (Figure 1 and Figure 2). Binding of the p21 peptide to PCNA involves an extensive set of interactions spanning the entire length of the peptide and burying 2,242 Å2 of total surface area at the intermolecular interface (57% on the peptide, 43% on PCNA).
The PCNA binding motif LIG_PCNA_PIPBox_1 reported for the region spanning residues 144 to 153, is located within the annotated region.
Cross-references: ELM:LIG_PCNA_PIPBox_1
PDB:1AXC
Protein: DP00016 (UniProt P38936)
Region: residues 139	to 143
Annotation: import into nucleus (Biological process)
Evidence: loss-of-function mutant phenotype evidence used in manual assertion
Supporting statements:
Here we identified the amino acids of p21 that are essential for its nuclear translocation and examined the consequences of their mutation. While the RKR140–142 sequence was essential for nuclear localization of p21, the KRK161–163 was not.
The annotated region is part of the TRG_NLS_Bipartite_1 motif associated with the nuclear localization signal (NLS), which has been described for the region spanning residues 142 to 158.
Cross-references: ELM:TRG_NLS_Bipartite_1
Protein: DP00016 (UniProt P38936)
Region: residues 143	to 147
Annotation: negative regulation of protein binding (Biological process)
Evidence: loss-of-function mutant phenotype evidence used in manual assertion
Supporting statements:
Simulated p21Cip1 phosphorylation at Thr 145 completely prevented p21Cip1-PCNA binding, whereas the nonphosphorylatable p21 T145A construct displayed no change in PCNA binding compared to p21 wt (Fig. 4A and B).
Therefore, we designed additional p21Cip1 constructs carrying a mutation to an unphosphorylatable (S146A) or to a phosphomimetic amino acid (S146D) at position 146. Coimmunoprecipitation experiments confirmed that in mammalian cells stimulation of p21Cip1 phosphorylation at serine 146 indeed decreases PCNA binding (Fig. 4D). However, phosphorylation of p21Cip1 at the Akt phosphorylation site Thr 145 exerts a more pronounced inhibition of complex formation with PCNA than Ser 146 phosphorylation (Fig. 4D). Taken together, Akt regulates p21Cip1-PCNA binding via specific phosphorylation of the Thr 145 residue.
Simulation of p21Cip1 phosphorylation at Thr 145 (T145D) reduced Cdk2 binding in endothelial cells by 59% ± 15% (Fig. ​5A) and affected complex formation with Cdk4 to a minor degree (Fig. ​5B), whereas both p21 wt and the T145A construct complexed with Cdk2 and Cdk4 to similar extents (Fig. ​5).
Through the analysis of a mimetic mutant, the authors show the phosphorylation of the 145 residue is a potent inhibitor of the binding to both PCNA and CDK2.
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: cyclin-dependent protein serine/threonine kinase inhibitor activity (Molecular function)
Evidence: in vivo protein kinase assay evidence used in manual assertion
Supporting statements:
Overexpression of p21Cip1 significantly reduced the kinase activity of Cdk2 (35% ± 6% of vector-transfected cells; P < 0.001). However, cells transfected with the Akt–phospho-mimetic p21Cip1 T145D construct displayed reduced inhibition of Cdk2 activity compared to cells overexpressing either p21 wt or the p21Cip1 T145A construct (Fig. 6C).
Protein: DP00016 (UniProt P38936)
Region: residues 143	to 147
Annotation: phosphorylation display site (Disorder function)
Evidence: in vivo protein kinase assay evidence used in manual assertion
Supporting statements:
To confirm that phosphorylation of p21Cip1 at Thr 145 occurs in vivo, we labeled intact cells with [32P]orthophosphate. Stimulation with serum induced the incorporation of 32P into endogenous p21Cip1 (Fig.3B).
These results indicate that serum induces phosphorylation of the peptide 144QTSMTDFYHSK154, which contains the Akt site Thr 145, in a PI3K-dependent manner. Taken together, the present data demonstrate that Akt interacts with and phosphorylates p21Cip1 in vitro and in vivo at Thr 145.
This annotation refers to the phosphorylation of the Thr 145 residue, included in the consensus motif for Akt-mediated phosphorylation MOD_PKB_1 that comprises the residues 140 to 148.
Cross-references: ELM:MOD_PKB_1
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: protein binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
p21 wt binds to PCNA as shown by coimmunoprecipitation studies with anti-myc antibodies for immunoprecipitation of myc-tagged p21 wt followed by Western blotting against endogenous PCNA (Fig. ​4A), as well as by immunoblot analysis of p21 bound to immunoprecipitates of endogenous PCNA (Fig. 4B).
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: PCNA-p21 complex (Cellular component)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
p21 wt binds to PCNA as shown by coimmunoprecipitation studies with anti-myc antibodies for immunoprecipitation of myc-tagged p21 wt followed by Western blotting against endogenous PCNA (Fig. ​4A), as well as by immunoblot analysis of p21 bound to immunoprecipitates of endogenous PCNA (Fig. 4B).
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
Annotation: protein serine/threonine kinase binding (Molecular function)
Evidence: co-immunoprecipitation evidence used in manual assertion
Supporting statements:
Simulation of p21Cip1 phosphorylation at Thr 145 (T145D) reduced Cdk2 binding in endothelial cells by 59% ± 15% (Fig. ​5A) and affected complex formation with Cdk4 to a minor degree (Fig. ​5B), whereas both p21 wt and the T145A construct complexed with Cdk2 and Cdk4 to similar extents (Fig. ​5).
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 49
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:DP00287r060
Protein: DP00016 (UniProt P38936)
Region: residues 1	to 164
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:DP00287r063
Protein: DP00017 (UniProt P49918)
Region: residues 1	to 316
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
The CD spectrum of p57 lacks the features that are characteristic of helical or β-sheet secondary structure and instead exhibits a strong negative band at approximately 203 nm that is reminiscent of an unfolded protein [Fig. 3(A)].
The CD spectra at 5 and 80°C are characteristic of an unfolded protein.
Protein: DP00017 (UniProt P49918)
Region: residues 1	to 316
Annotation: disorder (Structural state)
Evidence: fluorescence evidence used in manual assertion
Supporting statements:
Fluorescence emission spectrum of p57 (10 mM sodium phosphate, 50 mM NaCl, 1 mM DTT, pH 7.0, 30°C). The spectrum, collected with excitation at 280 nm, is characteristic of a solvent-exposed, unfolded protein.
The fluorescence emission spectrum exhibits a maximum at 350 nm [Fig. 3(B)], as expected for a solvent-exposed, unfolded protein.
Protein: DP00017 (UniProt P49918)
Region: residues 1	to 316
Annotation: disorder (Structural state)
Evidence: chromatography evidence used in manual assertion
Supporting statements:
The hydrodynamic radius of p57 determined with gel filtration is 32 Å (data not shown). A protein the size of p57 (316 amino acids) is expected to have hydrodynamic radii of 25 and 59 Å in globular and highly unfolded states, respectively.31 The hydrodynamic radius of p57 is approximately 1.3 times larger than expected for a monomeric, folded, globular protein, but significantly less than expected for a highly denatured protein.
Protein: DP00017 (UniProt P49918)
Region: residues 27	to 97
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
 Intrinsic Disorder of the p57 CDK-Inhibition Domain
The CD spectrum of p57ID is reminiscent of an unfolded protein, with a negative minimum at 202 nm [Fig. 5(A)].
CD analysis of p57ID (10 mM sodium phosphate, 50 mM NaCl, 1 mM DTT, pH 7.0). The CD spectra at 5 and 80°C are characteristic of an unfolded protein.
Protein: DP00017 (UniProt P49918)
Region: residues 27	to 97
Annotation: disorder (Structural state)
Evidence: fluorescence evidence used in manual assertion
Supporting statements:
Intrinsic Disorder of the p57 CDK-Inhibition Domain
The fluorescence spectrum of p27ID collected with an excitation wavelength of 280 nm exhibits an emission maximum at 350 nm [Fig. 5(B)], as expected for solvent-exposed aromatic residues in an unfolded protein.
Fluorescence emission spectrum of p57ID (10 mM sodium phosphate, 50 mM NaCl, 1 mM DTT, pH 7.0, 30 °C). The spectrum, collected with excitation at 280 nm, is characteristic of a solvent-exposed, unfolded protein.
Protein: DP00017 (UniProt P49918)
Region: residues 27	to 97
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Intrinsic Disorder of the p57 CDK-Inhibition Domain
The HN chemical shifts of p57ID span 7.8–8.5 ppm (Fig. 6), which are within the ranges expected of unfolded proteins.
1H-15N HSQC spectrum of p57ID (17 μM p57ID, 10 mM sodium phosphate, 50 mM NaCl, 1 mM DTT, pH 6.0, 30°C). Seventy resolved amide cross-peaks are observed from an expected total of 71 (76 residues minus Thr 1 and four Pro residues). The lack of HN chemical shift dispersion suggests that p57ID lacks stable secondary structure.
Protein: DP00017 (UniProt P49918)
Region: residues 27	to 97
Annotation: molecular function inhibitor activity (Molecular function)
Evidence: deletion mutation phenotypic evidence used in manual assertion
Supporting statements:
Similar to full-length p57, p57ID inhibits approximately 90% of the activity of cyclin A-CDK2 at a 1:1 molar ratio (Fig. 4), showing that the p57 CDK-inhibition domain is localized to the N-terminus, as expected from sequence comparisons with p21 and p27.
Protein: DP00017 (UniProt P49918)
Region: residues 1	to 61
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:DP00287r060
Protein: DP00017 (UniProt P49918)
Region: residues 1	to 316
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:DP00287r063
Protein: DP00018 (UniProt P46527)
Region: residues 38	to 59
Annotation: flexible linker (Disorder function)
Evidence: proton-based nuclear magnetic resonance evidence used in manual assertion
Supporting statements:
We call this segment the linker helix because it connects two domains, termed domain 1 and domain 2, that are conserved in the human CKIs (Fig. 1c).
Protein: DP00018 (UniProt P46527)
Region: residues 38	to 59
Annotation: disorder to order (Structural transition)
Evidence: proton-based nuclear magnetic resonance evidence used in manual assertion
Supporting statements:
We determined backbone 1HN, 13Cα and 15N chemical shifts for p27-KID (BioMagResBank entry 6112) and used secondary 13Cα chemical shift values (δΔ 13Cα)23 to identify those residues shown by CD to adopt helical secondary structure (Fig. 1a)24. Most residues show near random coil 13Cα chemical shift values (near zero δΔ 13Cα values). However, all residues between positions 37 and 59 show positive δΔ 13Cα values, consistent with α-helical secondary structure.
Protein: DP00018 (UniProt P46527)
Region: residues 105	to 198
Annotation: disorder (Structural state)
Evidence: proton-based nuclear magnetic resonance evidence used in manual assertion
Supporting statements:
p27 is comprised of an N-terminal domain (p27-KID, residues 22–104) that binds and inhibits Cdk/cyclin complexes and a C-terminal domain (residues 105–198, p27-C) that contains several sites of post-translational modification, including T187 (Fig. 1a).
We probed the structure and dynamics of the p27 C-terminus using NMR spectroscopy, first by studying the isolated C-terminal domain (p27-C) and second by studying isotope-labeled p27 bound to Cdk2/cyclin A. The 2D 1H-15N HSQC spectrum of 15N-labeled p27-C (Fig. 1b, green) exhibited the limited chemical shift dispersion that is typical of intrinsically unstructured proteins (IUPs). Further, secondary 13Cα chemical shift values (Δδ 13Cα) (Supplementary Fig. 1a) did not indicate the existence of highly populated secondary structure and {1H}-15N heteronuclear (hetNOE) values (Supplementary Fig. 1b) were almost exclusively negative, consistent with frequent, random amide group fluctuations on the high ps-low ns time scale. In addition, the chemical shift values of most amides within p27-C were very similar for the isolated domain and this domain in the context of 2H/15N-p27 bound to Cdk2/cyclin A (Fig. 1b, red). The structural independence of p27-C was confirmed by showing that resonances for p27-KID bound to Cdk2/cyclin A (Fig. 1c, blue) were recapitulated in the 2D 1H-15N TROSY spectrum of 2H/15N-p27/Cdk2/cyclin A (Fig. 1c, red). Together, these results indicated that p27-C in the p27/Cdk2/cyclin A complex is an independent, flexible domain that lacks secondary structure.
While the N-terminal KID of free p27 contained regions of transient structure, the C-terminal domain (p27-C, residues 105–198) lacked detectable secondary structure.
Here we have shown that the C-terminal domain of p27 adopts a disordered and quite highly extended conformation when p27 is bound to Cdk2/cyclin A.
Protein: DP00018 (UniProt P46527)
Region: residues 1	to 198
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
In contrast, the unbound p27 Cdk-inhibition domain is intrinsically disordered (natively unfolded) as shown by circular dichroism spectroscopy, lack of chemical-shift dispersion, and negative heteronuclear nuclear Overhauser effects. The intrinsic disorder is not due to the excision of the Cdk-inhibition domain from p27, since circular dichroism spectra of the full-length protein are also indicative of a largely unfolded protein.
Intrinsic Structural Disorder of p27. The full-length p27 used here inhibits histone H1 phosphorylation by cyclin A-Cdk2 (Figure 2). The CD spectrum of full-length human p27 is characteristic of an unfolded protein (Figure 3). The minimum at 200 nm is indicative of an unfolded conformation, and the weak negative shoulder at 222 nm suggests the absence of significant amounts of helix or β-strand. The helix content is estimated at 2 or 6% from [θ]222 or CDPro, respectively. The results indicate that full-length p27, while active as a Cdk inhibitor, is largely unfolded.
The negative ellipticity at 200 nm and the lack of significant ellipticity at 216 nm and above are typical of the CD spectra of unfolded proteins.