Cryo-EM data collection, refinement and validation statistics

Similar documents
SUPPLEMENTARY INFORMATION

Structure and RNA-binding properties. of the Not1 Not2 Not5 module of the yeast Ccr4 Not complex

Structure of the quaternary complex between SRP, SR, and translocon bound to the translating ribosome

Nature Structural & Molecular Biology: doi: /nsmb Supplementary Figure 1

Table S1. Overview of used PDZK1 constructs and their binding affinities to peptides. Related to figure 1.

Supplementary Figure 1 Crystal contacts in COP apo structure (PDB code 3S0R)

Supplementary Figure 1. Biochemical and sequence alignment analyses the

SUPPLEMENTARY INFORMATION

Full wwpdb/emdatabank EM Map/Model Validation Report i

Nature Structural & Molecular Biology: doi: /nsmb Supplementary Figure 1

SUPPLEMENTARY INFORMATION

Supplementary Materials for

SUPPLEMENTARY INFORMATION

SUPPLEMENTARY INFORMATION

SUPPLEMENTARY INFORMATION

type GroEL-GroES complex. Crystals were grown in buffer D (100 mm HEPES, ph 7.5,

Purification, SDS-PAGE and cryo-em characterization of the MCM hexamer and Cdt1 MCM heptamer samples.

SUPPLEMENTARY INFORMATION

SUPPLEMENTARY INFORMATION

Supporting Protocol This protocol describes the construction and the force-field parameters of the non-standard residue for the Ag + -site using CNS

Spliceosome and Localization of Its Catalytic Core

Table 1. Crystallographic data collection, phasing and refinement statistics. Native Hg soaked Mn soaked 1 Mn soaked 2

SUPPLEMENTARY INFORMATION

Supplementary figure 1 Application of tmfret in LeuT. (a) To assess the feasibility of using tmfret for distance-dependent measurements in LeuT, a

Supplemental Information. The Mitochondrial Fission Receptor MiD51. Requires ADP as a Cofactor

SUPPLEMENTARY INFORMATION

York University School of Medicine, 550 First Avenue, MSB 599, New York, New York 10016, USA.

RNA Polymerase I Contains a TFIIF-Related DNA-Binding Subcomplex

Supplementary Figures

Acta Crystallographica Section D

Supplementary figure 1. Comparison of unbound ogm-csf and ogm-csf as captured in the GIF:GM-CSF complex. Alignment of two copies of unbound ovine

Supplementary Figures

of the Guanine Nucleotide Exchange Factor FARP2

Nature Structural and Molecular Biology: doi: /nsmb Supplementary Figure 1

SUPPLEMENTARY INFORMATION

Supplemental Data SUPPLEMENTAL FIGURES

Full wwpdb X-ray Structure Validation Report i

SUPPLEMENTARY INFORMATION

SI Text S1 Solution Scattering Data Collection and Analysis. SI references

Expanded View Figures

Diphthamide biosynthesis requires a radical iron-sulfur enzyme. Pennsylvania State University, University Park, Pennsylvania 16802, USA

SUPPLEMENTARY INFORMATION

SUPPLEMENTARY INFORMATION

Nature Structural & Molecular Biology: doi: /nsmb Supplementary Figure 1

Lipid Regulated Intramolecular Conformational Dynamics of SNARE-Protein Ykt6

Supplemental Information. Expanded Coverage of the 26S Proteasome. Conformational Landscape Reveals. Mechanisms of Peptidase Gating

Supporting Information

Supplementary Information: Long range allosteric regulation of the human 26S proteasome by 20S proteasome-targeting cancer drugs

THE CRYSTAL STRUCTURE OF THE SGT1-SKP1 COMPLEX: THE LINK BETWEEN

Acta Crystallographica Section D

Supplemental Information. Molecular Basis of Spectral Diversity. in Near-Infrared Phytochrome-Based. Fluorescent Proteins

Structure and mechanism of an intramembrane liponucleotide synthetase central for phospholipid biosynthesis

Supplementary Information. The protease GtgE from Salmonella exclusively targets. inactive Rab GTPases

Structural characterization of NiV N 0 P in solution and in crystal.

SUPPLEMENTARY INFORMATION

Nature Structural and Molecular Biology: doi: /nsmb.2938

Supplementary Figure 1. Aligned sequences of yeast IDH1 (top) and IDH2 (bottom) with isocitrate

Full wwpdb X-ray Structure Validation Report i

Plasmid Relevant features Source. W18N_D20N and TrXE-W18N_D20N-anti

Supplementary Information. Recognition of the pre-mirna structure by Drosophila Dicer-1

Nature Structural & Molecular Biology: doi: /nsmb Supplementary Figure 1

Full wwpdb X-ray Structure Validation Report i


Homology Modeling (Comparative Structure Modeling) GBCB 5874: Problem Solving in GBCB

Supporting Information

SUPPLEMENTARY INFORMATION

Supplementary Figure 1

SUPPLEMENTARY INFORMATION. doi: /nature07461

Tools for Cryo-EM Map Fitting. Paul Emsley MRC Laboratory of Molecular Biology

Full wwpdb X-ray Structure Validation Report i

Protein structure analysis. Risto Laakso 10th January 2005

Full wwpdb X-ray Structure Validation Report i

Structural insights into Aspergillus fumigatus lectin specificity - AFL binding sites are functionally non-equivalent

Full wwpdb X-ray Structure Validation Report i

Full wwpdb X-ray Structure Validation Report i

Enhancing hydrogen production of microalgae by redirecting electrons from photosystem I to hydrogenase

Nature Structural and Molecular Biology: doi: /nsmb.2783

Full wwpdb X-ray Structure Validation Report i

Nature Structural & Molecular Biology doi: /nsmb Supplementary Figure 1. CRBN binding assay with thalidomide enantiomers.

Introduction to Comparative Protein Modeling. Chapter 4 Part I

Supplementary text for the section Interactions conserved across species: can one select the conserved interactions?

SUPPLEMENTARY INFORMATION

According to the manufacture s direction (Pierce), RNA and DNA

Full wwpdb X-ray Structure Validation Report i

Full wwpdb X-ray Structure Validation Report i

Orientational degeneracy in the presence of one alignment tensor.

Nature Structural & Molecular Biology: doi: /nsmb Supplementary Figure 1

SUPPLEMENTARY INFORMATION

Automated identification of functional dynamic contact networks from X-ray crystallography

Stabilizing the CH2 domain of an Antibody by Engineering in an Enhanced Aromatic Sequon

Supplementary Figure 1. SDS-PAGE analysis of GFP oligomer variants with different linkers. Oligomer mixtures were applied to a PAGE gel containing

SUPPLEMENTARY INFORMATION

SUPPLEMENTARY FIGURES

Nature Structural and Molecular Biology: doi: /nsmb Supplementary Figure 1. Definition and assessment of ciap1 constructs.

Supplementary Materials for

SUPPLEMENTARY INFORMATION

Supplementary Information

SUPPLEMENTARY INFORMATION

The copper active site in CBM33 polysaccharide oxygenases

Structure and Function of Neisseria gonorrhoeae MtrF Illuminates a Class of Antimetabolite Efflux Pumps

Transcription:

1 Table S1 Cryo-EM data collection, refinement and validation statistics Data collection and processing CPSF-160 WDR33 (EMDB-7114) (PDB 6BM0) CPSF-160 WDR33 (EMDB-7113) (PDB 6BLY) CPSF-160 WDR33 CPSF-30 PAS RNA (EMDB-7112) (PDB 6BLL) Magnification 22,500 22,500 22,500 Voltage (kv) 300 300 300 Electron exposure (e /Å 2 ) 47 70 70 Defocus range (μm) 1.3~2.7 1.2~2.5 1.2~2.5 Pixel size (Å) 1.30 1.07 1.07 Symmetry imposed C1 C1 C1 Image stacks (no.) 1625 2486 2486 Initial particle images (no.) 372,707 1,144,122 1,144,122 Final particle images (no.) 205,373 456,310 173,632 Map resolution (Å) FSC threshold 3.78 0.143 3.36 0.143 3.42 0.143 Map sharpening B-factor (Å 2 ) 180 186 157 Refinement Number of protein residues 1,538 1,540 1,669 Number of RNA nucleotides Number of metal ions 0 0 0 0 7 3 Number of atoms 12,244 12,263 13,435 R.m.s. deviations Bond lengths (Å) Bond angles ( ) 0.008 1.05 0.008 0.98 0.010 1.02 PDB validation Clash score Poor rotamers (%) Ramachandran plot Favored (%) Allowed (%) Disallowed (%) 18 0.52 91.06 7.81 1.13 7 0.22 91.20 8.47 0.33 10 0.28 90.48 9.33 0.18

Fig. S1. Image processing of the CPSF-160 WDR33 complex. (A). Area of an image of the complex in negative stain. Scale bar: 50 nm. (B). Selected 2D class averages obtained with the ISAC algorithm. Side length of individual averages: 19 nm. (C). The initial model generated with the VIPER algorithm using the ISAC averages obtained for the negatively stained complex. (D). Area of an image of the vitrified complex. Some particles are circled. Scale bar: 50 nm. (E). Selected 2D class averages obtained with RELION-2. Side length of individual averages: 21 nm. (F). Image-processing workflow for 3D classification and refinement in RELION-2 that resulted in a density map at 3.8 Å resolution. See Methods for details. 2

Fig. S2. Image processing of the CPSF-160 WDR33 CPSF-30 PAS RNA complex. (A). Area of a cryo-em image of the vitrified complex. Some particles are circled. Scale bar: 50 nm. (B). Selected 2D class averages obtained with RELION-2. Side length of individual averages: 21 nm. (C). Image-processing workflow for 3D classification and refinement in RELION-2 that resulted in a density map at 3.42 Å resolution. See Methods for details. 3

Fig. S3. FSC curves and local densities. (A). Gold-standard FSC curves calculated between independently refined half maps for (1) the density map of the CPSF-160 WDR33 complex (labeled 160-33, yellow), (2) the density map obtained with the CPSF-160 WDR33 CPSF-30 PAS RNA sample by combining five classes after the first 3D classification (Map 1 in Supplementary Fig. 2) (red), and (3) the density map obtained with the CPSF-160 WDR33 CPSF-30 PAS RNA sample by combining five classes after the second 3D classification (Map 2 in Supplementary Fig. 2) (green). (B). Cross-validation FSC curves: green, refined model versus half map 1 used for refinement (work map); yellow, refined model versus half map 2 not used for refinement (free map); red, refined model versus the combined final map. The similarity of the work and free curves suggests no substantial over-fitting. (C). Representative cryo-em densities for Map2 obtained with the CPSF-160 WDR33 CPSF-30 PAS RNA sample. (D). Two views of the cryo-em map of the quaternary complex, segmented and colored as in panel a of Fig. 1. The views are related by 90 rotation around the vertical axis. (E). Two views of the cryo-em map of the binary complex at 3.8 Å resolution. 4

5 Fig. S4. Sequence alignment of selected WDR33 homologs. The secondary structure elements in the human WDR33 structure are shown. Residues in contact with CPSF-160, CPSF-30, and PAS RNA are indicated with purple, green, and orange dots, respectively. The boundaries of the WD40 domain are indicated. Only residues 1-451 of human WDR33 are included in the alignment. Hs: Homo sapiens, Mm: Mus musculus, Xt: Xenopus tropicalis; Dr: Danio rerio, Dm: Drosophila melanogaster.

Fig. S5. Overlay of the structures of the quaternary and binary complexes. (A). Overlay of the structures of the CPSF- 160 WDR33 CPSF-30 PAS RNA quaternary complex (in color) and the CPSF-160 WDR33 binary complex (gray). (B). Close-up view of the N-terminal segment of WDR33 containing residues 43-54 (cyan). This segment is disordered in the binary complex (gray). 6

Fig. S6. Sequence alignment of selected CPSF-30 homologs. The secondary structure elements in the human CPSF- 30 structure are shown. Residues in contact with CPSF-160, WDR33, and PAS RNA are indicated with purple, light blue, and orange dots, respectively. The boundaries of ZF1-ZF3 are indicated. Hs: Homo sapiens, Mm: Mus musculus, Xt: Xenopus tropicalis; Dr: Danio rerio, Dm: Drosophila melanogaster. 7

Fig. S7. Comparison of the RNA-binding mode in ZF2 of CPSF-30 with that in other CCCH zinc fingers. (A). Overlay of the structures of CPSF-30 ZF2 and Nab2 ZF6 (PDB ID 5L2L). (B). Overlay of the structures of CPSF- 30 ZF2 and MBNL1 ZF3 (PDB ID 3D2S). (C). Overlay of the structures of CPSF-30 ZF2 and TIS11d ZF1 (PDB ID 1RGO). 8

Fig. S8. Gel filtration profiles for purified mpsf proteins used for biochemical studies. Gel filtration profiles for the CPSF-160 WDR33 CPSF-30 ternary complex (red), the CPSF-160 WDR33 binary complex (blue), CPSF-160 alone (cyan), MBP-CPSF-30 fusion protein (magenta), and a mixture of CPSF-160 and MBP-CPSF-30 (orange) are shown. Inset: SDS-PAGE gel of the protein samples. The second peak for the ternary complex contains MBP and excess CPSF-30 (lane 5). 9

Fig. S9. Structural similarity between the CPSF-160 WDR33 complex and the DDB1 DDB2 complex. (A). Overlay of the structures of the CPSF-160 WDR33 CPSF-30 PAS RNA quaternary complex (in color) with that of the DDB1 DDB2 dsdna complex (gray, PDB ID 3EI1). The overlay is between CPSF-160 and DDB1 only. (B). Overlay of the structures of the CPSF-160 WDR33 CPSF-30 PAS RNA quaternary complex (in color) with that of the DDB1 CUL4A RBX1 SV5-V complex (gray, PDB ID 2HYE). The overlay is based on BPA and BPC of CPSF-160. The protein bound between BPA and BPC of DDB1 (SV5-V) does not contain a b-propeller. The bottom face of BPB mediates interaction with CUL4A. 10

2024 © sciencedocbox.com Privacy Policy | Terms of Service | Feedback