LiteFold/DisProt huggingface dataset:
Protein: DP03326 (UniProt P39286)
Region: residues 1	to 34
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
A major species-specific feature of RsgA from E. coli is that it possesses an additional N-terminal extension, which is disordered in the crystal structure of RsgA (19) and therefore absent in our atomic model as well (missing residues 1–34). Because of its flexible nature, we were only able to locate partial densities for this N-terminal extension.
 The first 34 residues were cut from the model because of the disorderness in the template.
The RsgA N-terminal extension is disordered in the crystal structure, and thus absent in the atomic model. Partially resolved densities for the N extension show that it interacts with h18, h34, and S3.
Cross-references: PDB:2YKR
EMDB:1884
Protein: DP03326 (UniProt P39286)
Region: residues 1	to 34
Annotation: rRNA binding (Molecular function)
Evidence: cryogenic electron microscopy evidence used in manual assertion
Supporting statements:
A major species-specific feature of RsgA from E. coli is that it possesses an additional N-terminal extension, which is disordered in the crystal structure of RsgA (19) and therefore absent in our atomic model as well (missing residues 1–34). Because of its flexible nature, we were only able to locate partial densities for this N-terminal extension. Nevertheless, the map clearly indicates that this extension interacts at least with helix 18 (loop 530) of the body, and helix 34 and S3 of the head (Fig. S2), consistent with the fact that the first 20 N-terminal residues are required for the 30S subunit binding.
The RsgA N-terminal extension is disordered in the crystal structure, and thus absent in the atomic model. Partially resolved densities for the N extension show that it interacts with h18, h34, and S3.
Cross-references: PDB:2YKR
EMDB:1884
Protein: DP03327 (UniProt P05055)
Region: residues 553	to 612
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
The C-terminal KH and S1 domains (Fig. 1A, residues 566–734) were not visible, containing only broken densities in the electron density maps. These KH and S1 domains were likely disordered as a whole in the crystal structure since they were connected to the second RNase PH domain by a long flexible loop.
The C-terminal KH/S1 domain (marked by dashed circle) was disordered and thus not visible in the crystal structure.
The IDR was separated according to the protein domain composition. This corresponds to the domain "KH".
Cross-references: PDB:3CDI
Protein: DP03327 (UniProt P05055)
Region: residues 238	to 298
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
These results suggest that the structure of DKH/S1 was less compact than full-length PNPase.
To further confirm this observation, the sizes of central channel of full-length PNPase and DKH/S1 mutant were calculated by Hole (Smart et al. 1993). To ensure a fair comparison, disordered side chains or side chains with different conformations in the two structures were removed for the calculation, including the side chains of 105–107, 261–321, 394, and 409–418.
 The IDR characterized in the publication and spanning residues 261–321 corresponds to region 238-298 of the amino acid sequence.
Cross-references: PDB:3CDJ
Protein: DP03327 (UniProt P05055)
Region: residues 386	to 395
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
These results suggest that the structure of DKH/S1 was less compact than full-length PNPase.
To further confirm this observation, the sizes of central channel of full-length PNPase and DKH/S1 mutant were calculated by Hole (Smart et al. 1993). To ensure a fair comparison, disordered side chains or side chains with different conformations in the two structures were removed for the calculation, including the side chains of 105–107, 261–321, 394, and 409–418.
 The IDR characterized in the publication and spanning residues 409-418 corresponds to region 386-395 of the amino acid sequence.
Cross-references: PDB:3CDJ
Protein: DP03327 (UniProt P05055)
Region: residues 543	to 711
Annotation: RNA binding (Molecular function)
Evidence: electrophoretic mobility shift assay evidence used in manual assertion
Supporting statements:
We purified full-length PNPase (Fig. 1A, residues 17–734) and the KH/S1 domain-truncated mutant (Fig. 1A, DKH/S1, residues 17–570), and compared their RNA binding activities by electrophoresis mobility shift assays, using a 20-mer and an 8-mer single-stranded RNA as substrates. The EMSA (Fig. 1B) showed that the full-length PNPase bound 20-mer and 8-mer RNA at concentrations >4 nM, whereas the truncated mutant DKH/S1 bound 20-mer and 8-mer RNA at concentrations >z20 nM.
The KH/S1 domain pair is responsible for direct interaction with RNA.
Our biochemical, circular dichroism, dynamic light scat- tering, and structural results consistently suggest that the KH/S1 domains are involved not only in RNA binding but also in the formation of a compact trimer with a more constricted central channel
Protein: DP03327 (UniProt P05055)
Region: residues 543	to 711
Annotation: molecular function regulator (Molecular function)
Evidence: enzymatic activity assay evidence used in manual assertion
Supporting statements:
We then tested the exoribonucleolytic activity of full- length PNPase and DKH/S1 mutant in time course activity assays. The full-length PNPase digested both 20-mer and 8- mer RNA into small oligonucleotides, as shown in Figure 1C.
These results imply that the KH/S1 domain is involved not only in RNA binding, but also in modulation of enzyme activity in an unknown way.
RNase activities of full-length PNPase and DKH/S1 mutant were assayed by incubation of enzymes, respectively, with 8-mer and 20-mer single-stranded RNAs in time course experiments under the conditions described in Materials and Methods. The left panel shows that DKH/S1 had lower activities in cleaving 20-mer ssRNAs compared to the full-length PNPase. The right panel shows that DKH/S1 cannot degrade shorter 8-mer ssRNAs.
Cross-references: PDB:3CDI
Protein: DP03327 (UniProt P05055)
Region: residues 543	to 711
Annotation: RNA binding (Molecular function)
Evidence: circular dichroism evidence used in manual assertion
Supporting statements:
This result showed that the truncated PNPase, without the KH/S1 domain, was melted at lower temperatures and dissociated into monomers in less acidic solutions. Therefore, we conclude that the KH and S1 domains contribute to the formation of a more stable trimeric PNPase.
The KH/S1 domain helps PNPase to form a more stable compact trimer. Our biochemical, circular dichroism, dynamic light scattering, and structural results consistently suggest that the KH/S1 domains are involved not only in RNA binding but also in the formation of a compact trimer with a more constricted central channel.
We conclude that the KH/S1 domains and the upper neck in the channel recruit RNA into the channel, and the lower neck keeps hold of the RNA as it guides it into the PNPase active site for processive degradation. Constrictions of the appropriate size at the necks in the central channel of exosomes and PNPases play a crucial role in RNA binding and processive degradation.
Cross-references: PDB:3CDI
Protein: DP03327 (UniProt P05055)
Region: residues 543	to 711
Annotation: RNA binding (Molecular function)
Evidence: dynamic light scattering assay evidence used in manual assertion
Supporting statements:
This result showed that the truncated PNPase, without the KH/S1 domain, was melted at lower temperatures and dissociated into monomers in less acidic solutions. Therefore, we conclude that the KH and S1 domains contribute to the formation of a more stable trimeric PNPase.
The KH/S1 domain helps PNPase to form a more stable compact trimer. Our biochemical, circular dichroism, dynamic light scattering, and structural results consistently suggest that the KH/S1 domains are involved not only in RNA binding but also in the formation of a compact trimer with a more constricted central channel.
We conclude that the KH/S1 domains and the upper neck in the channel recruit RNA into the channel, and the lower neck keeps hold of the RNA as it guides it into the PNPase active site for processive degradation. Constrictions of the appropriate size at the necks in the central channel of exosomes and PNPases play a crucial role in RNA binding and processive degradation.
Cross-references: PDB:3CDI
Protein: DP03327 (UniProt P05055)
Region: residues 75	to 84
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
This region is not described in the publication, although it corresponds to a region with missing electron density in the PDB structure. 
Cross-references: PDB:3CDJ
Protein: DP03327 (UniProt P05055)
Region: residues 359	to 370
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
This region is not described in the publication, although it corresponds to a region with missing electron density in the PDB structure. 
Cross-references: PDB:3CDJ
Protein: DP03327 (UniProt P05055)
Region: residues 622	to 690
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
The C-terminal KH and S1 domains (Fig. 1A, residues 566–734) were not visible, containing only broken densities in the electron density maps. These KH and S1 domains were likely disordered as a whole in the crystal structure since they were connected to the second RNase PH domain by a long flexible loop.
The C-terminal KH/S1 domain (marked by dashed circle) was disordered and thus not visible in the crystal structure.
The IDR was separated according to the protein domain composition. This corresponds to the domain "S1".
Cross-references: PDB:3CDI
Protein: DP03329 (UniProt P0AG67)
Region: residues 73	to 86
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
Deep classification of cryo-EM data reveals that the N-terminal helix of S1 acts as an anchor (Fig. 2C), whereas the rest of S1 is extremely dynamic (Movie 1).
However, continuous density was visible only in one or two classes and was not of sufficient quality to dock S1 domains. Overall, this unbiased global classification was useful in that it revealed S1 binding site, however the separation of classes occurred on too many different features of the 70S ribosome and the classes did not reveal domain distribution for S1.
We compared four approaches for deep classification of a ribosome dataset to identify conformational states of a highly dynamic protein S1.
In the next step, a spherical mask around the region of interest, while masking out the large macromolecule using a 3D mask (negative masking), identifies different classes of the mobile protein in the region of interest.
This work highlights that while critical mechanistic insights can be gleaned at the ribosome core of an ensemble of high-resolution structures (Loveland et al., 2017), the same data set does not provide high resolution for a flexible peripheral protein even after exhaustive classification into up to 48 classes.
By contrast, domain 1 (D1), which is connected with the N-terminal helix via a 10-aa linker, is highly dynamic and adopts multiple positions, which differ by up to 50 Å.
While the N-terminal helix is relatively static, the rest of domain 1 is highly dynamic and interacts with S2 in many, but not all, classes.
Cross-references: PDB:6BU8
EMDB:EMD-7289
Protein: DP03329 (UniProt P0AG67)
Region: residues 180	to 557
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
Deep classification of cryo-EM data reveals that the N-terminal helix of S1 acts as an anchor (Fig. 2C), whereas the rest of S1 is extremely dynamic (Movie 1).
However, continuous density was visible only in one or two classes and was not of sufficient quality to dock S1 domains. Overall, this unbiased global classification was useful in that it revealed S1 binding site, however the separation of classes occurred on too many different features of the 70S ribosome and the classes did not reveal domain distribution for S1.
We compared four approaches for deep classification of a ribosome dataset to identify conformational states of a highly dynamic protein S1.
In the next step, a spherical mask around the region of interest, while masking out the large macromolecule using a 3D mask (negative masking), identifies different classes of the mobile protein in the region of interest.
This work highlights that while critical mechanistic insights can be gleaned at the ribosome core of an ensemble of high-resolution structures (Loveland et al., 2017), the same data set does not provide high resolution for a flexible peripheral protein even after exhaustive classification into up to 48 classes.
Cross-references: PDB:6BU8
EMDB:EMD-7289
Protein: DP03329 (UniProt P0AG67)
Region: residues 73	to 86
Annotation: protein binding (Molecular function)
Evidence: cryogenic electron microscopy evidence used in manual assertion
Supporting statements:
While the N-terminal helix is relatively static, the rest of domain 1 is highly dynamic and interacts with S2 in many, but not all, classes.
Cross-references: PDB:6BU8
EMDB:EMD-7289
Protein: DP03329 (UniProt P0AG67)
Region: residues 194	to 262
Annotation: mRNA binding (Molecular function)
Evidence: cryogenic electron microscopy evidence used in manual assertion
Supporting statements:
To visualize the region that the dynamic domains of S1 sample, we superimposed all the modeled domains from 15 classes (Fig. 5E). This superposition revealed that S1 excursions completely cover the 5′ tail of the mRNA upstream of the Shine-Dalgarno helix, rather than adopting preferred conformations that would leave a well-defined path for mRNA exit. Thus, S1 dynamics likely increases the probability of interactions with various mRNA structures near the mRNA exit, allowing S1 to facilitate mRNA association with the 30S subunit during initiation.
 It is possible that the density corresponds to the multiple, interchangeable domains, yielding poor resolution. We have placed D3 in the density on the basis of connectivity with D2, crosslinking of S1 with the 30S subunit (Lauber et al., 2012) and NMR studies that identified the face of S1 domains that interacts with RNA (Aliprandi et al., 2008). One prominent position is near the mRNA tail (Fig. 4A–C). Our finding of one rather than multiple S1 domain bound to the four 5′ mRNA residues is consistent with the observation that S1 domains bind on average 2.5 nucleotides of RNA.
Our cryo-EM maps are also consistent with binding of the C-terminal domains of S1 to the 5′ end of the mRNA and the 3′ end of 16S rRNA at the mRNA exit channel.
The region boundaries are defined by those corresponding to the C-terminal domains of S1.
Cross-references: PDB:6BU8
EMDB:EMD-7289
Protein: DP03329 (UniProt P0AG67)
Region: residues 441	to 557
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
The bS1 protein comprises six structurally related oligosaccharide-oligonucleotide binding (OB-fold) domains (D1–D6), of which we observe cryo-EM density for D1 to D5 in the hibernating 70S (Fig. 3c,d and Supplementary Fig. 5a,b).
Cross-references: PDB:6H4N
EMDB:EMD-0137
Protein: DP03329 (UniProt P0AG67)
Region: residues 177	to 263
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
With the exception of bS1-D3, which was highly flexible and poorly ordered (Supplementary Fig. 5b), molecular models could be unambiguously fitted for D2, D4 and D5 of bS1 (Fig. 3e and Supplementary Fig. 5e–j).
Cross-references: PDB:6H4N
EMDB:EMD-0137
Protein: DP03330 (UniProt P0A705)
Region: residues 1	to 381
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
The IF2 domain IV/ A76-fMet module is linked by flexible regions to the core of IF2 and the body of the tRNA, respectively, to uncouple the motion and to buffer the rotational movement of the 30S subunit. On the IF2 side, a flexible loop connects domain IV that extends from helix 12 of domain III.
The structure of poorly resolved, flexible loops in IF2 was predicted by FragFit, a fragment-based tool for modeling of missing protein segments into cryo-EM density maps (see section below).
Helix 12 of IF2 fades out into a flexible loop that connects domain III and IV while in eIF5b both domains are connected by a more rigid α-helix explaining the different modes of motion employed for repositioning of the initiator tRNA.
The same region is also present in the structure with PDB code "3JCJ".
Cross-references: PDB:3JCN
PDB:3JCJ
Protein: DP03330 (UniProt P0A705)
Region: residues 1	to 381
Annotation: disorder (Structural state)
Evidence: cryogenic electron microscopy-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
In both 30S IC-1 and IC-2 maps, a second larger density (purple density in Figure 1B and E) is located at the entrance to the mRNA channel suitable to account for parts of the N1 and N2 domains which were shown by NMR to consist of a small ordered fold and a larger unstructured region (37).
In panels B–C and E–F the star indicates density that we attribute to the G1 domain of IF2 while the dark purple density located on the back of the subunit near the mRNA entrance channel is attributed to the N1/N2 domain of IF2.
The same region is also present in the structure with PDB code "5ME1".
Cross-references: PDB:5ME0
PDB:5ME1
EMDB:EMD-3494
Protein: DP03330 (UniProt P0A705)
Region: residues 1	to 294
Annotation: molecular condensate scaffold activity (Molecular function)
Evidence: loss-of-function mutant phenotype evidence used in manual assertion
Supporting statements:
We observed IF2ΔNTD showed a notable reduction in IF2 droplets compared to the full length IF2 protein, suggesting the N-terminal IDR promotes IF2 phase separation. We also tested the β-isoform lacking the first 158 amino acids and found that the loss of this region leads to a robust loss in phase separation (Fig S5).
Protein: DP03331 (UniProt P0A8N3)
Region: residues 445	to 456
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
No electron density is present for the 10 N-terminal amino acids, the 2 C-terminal amino acids, for residue 269 and the regions enclosed between residues 154-160, 215-217, and 444-455.
The two loops (215-217 and 444-455) which are disordered in the uncomplexed LysS structure are indicated in magenta.
Whereas in the unliganded enzyme the regions between residues 215-217 and 444-455 are completely disordered, in the lysine-bound structure both segments of polypeptide chain show well-defined electron density.
The two loops that are disordered in the unliganded structure are colored in red. A space filling model of the lysine is shown in yellow. (C) A close up of the active site showing a stereo superposition of the lysine-bound (green) and unliganded (blue) LysS. In the unliganded structure the loops that are disordered (residues 215-218 and 444-455) have been modeled for the sake of clarity and are shown in red.
Cross-references: PDB:1BBW
Protein: DP03331 (UniProt P0A8N3)
Region: residues 445	to 456
Annotation: disorder to order (Structural transition)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
The two loops (215-217 and 444-455) which are disordered in the uncomplexed LysS structure are indicated in magenta.
Whereas in the unliganded enzyme the regions between residues 215-217 and 444-455 are completely disordered, in the lysine-bound structure both segments of polypeptide chain show well-defined electron density.
The two loops that are disordered in the unliganded structure are colored in red. A space filling model of the lysine is shown in yellow. (C) A close up of the active site showing a stereo superposition of the lysine-bound (green) and unliganded (blue) LysS. In the unliganded structure the loops that are disordered (residues 215-218 and 444-455) have been modeled for the sake of clarity and are shown in red.
Cross-references: PDB:1BBU
Protein: DP03332 (UniProt P0A3B2)
Region: residues 32	to 55
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
Disordered regions are indicated with dashed lines. B, sequence alignment of EF-G, LepA, BipA, and C-terminal fragment of BipA with domains color-coded as in A. Residue number below the sequence indicates residue N-terminal to the boundary between domains. The beginning and end of disordered regions are indicated with the residue number of the last ordered residue on the top of the sequence. Disordered regions are displayed as an array of thin boxes.
Cross-references: PDB:4ZCI
Protein: DP03332 (UniProt P0A3B2)
Region: residues 540	to 555
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
Disordered regions are indicated with dashed lines. B, sequence alignment of EF-G, LepA, BipA, and C-terminal fragment of BipA with domains color-coded as in A. Residue number below the sequence indicates residue N-terminal to the boundary between domains. The beginning and end of disordered regions are indicated with the residue number of the last ordered residue on the top of the sequence. Disordered regions are displayed as an array of thin boxes.
 A long, polar region (residues 540 –555) that extends from the distal end of the C-terminal domain is disordered in the crystal structure.
To evaluate the significance of the bound cations and to resolve the long, disordered, polar region, we set out to crystallize a C-terminal fragment of BipA consisting of domains III and V and the C-terminal domain in the presence of magnesium ions (Fig. 2A).
Cross-references: PDB:4ZCI
Protein: DP03332 (UniProt P0A3B2)
Region: residues 32	to 55
Annotation: disorder (Structural state)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
The “switch 1” region of BipA (residues 42–65), whose position is affected by the γ-phosphate of GTP in other GTPases, is disordered.
The IDR characterized in the publication with residues 42-65, corresponds to region 32-55 of the amino acid sequence.
Cross-references: PDB:4ZCL
Protein: DP03332 (UniProt P0A3B2)
Region: residues 540	to 555
Annotation: disorder to order (Structural transition)
Evidence: X-ray crystallography-based structural model with missing residue coordinates used in manual assertion
Supporting statements:
In addition, nearly all of the long, basic, disordered region is now structured, and the position of the [Co(NH3 )6 ]3+ ion in the full-length protein structure has been replaced by two proximal magnesium ions.
This evidence comes from a protein fragment.
Cross-references: PDB:4ZCK