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- EMDB-62369: Focused refinement map of human PNPase in open form -

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Open data


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Basic information

Entry
Database: EMDB / ID: EMD-62369
TitleFocused refinement map of human PNPase in open form
Map data
Sample
  • Complex: Human PNPase in open form
    • Protein or peptide: Human polynucleotide phosphorylase (hPNPase)
Keywords3'-to-5' exoribonuclease / RNA degradation / RNA import / mitochondria / RNA BINDING PROTEIN
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.72 Å
AuthorsLi YC / Yuan HS
Funding support Taiwan, 1 items
OrganizationGrant numberCountry
Academia Sinica (Taiwan) Taiwan
CitationJournal: Nucleic Acids Res / Year: 2025
Title: Structural insights into human PNPase in health and disease.
Authors: Yi-Ching Li / Chun-Hsiung Wang / Malay Patra / Yi-Ping Chen / Wei-Zen Yang / Hanna S Yuan /
Abstract: Human polynucleotide phosphorylase (hPNPase) is a 3'-to-5' exoribonuclease located in mitochondria, where it plays crucial roles in RNA degradation and RNA import. Mutations in hPNPase can impair ...Human polynucleotide phosphorylase (hPNPase) is a 3'-to-5' exoribonuclease located in mitochondria, where it plays crucial roles in RNA degradation and RNA import. Mutations in hPNPase can impair these functions, leading to various mitochondrial dysfunctions and diseases. However, the mechanisms by which hPNPase switches between its roles as an RNA-degrading enzyme and an RNA carrier, as well as how disease-associated mutations may affect these distinct functions, remain unclear. In this study, we present cryo-electron microscopy structures of hPNPase, highlighting the flexibility of its S1 domains, which cap the ring-like RNA-degradation chamber and shift between two distinctive open and closed conformations. We further demonstrate by small-angle X-ray scattering and biochemical analyses that the disease-associated mutations P467S and G499R impair hPNPase's stem-loop RNA-binding and degradation activities by limiting the S1 domain's ability to transition from an open to closed state. Conversely, the D713Y mutation, located within the S1 domain, does not affect the RNA-binding affinity of hPNPase, but diminishes its interaction with Suv3 helicase for cooperative degradation of structured RNA. Collectively, these findings underscore the critical role of S1 domain mobility in capturing structured RNA for degradation and import, as well as its involvement in mitochondrial degradosome assembly. Our study thereby reveals the molecular mechanism of hPNPase in RNA binding and degradation, and the multiple molecular defects that could be induced by disease-linked mutations in hPNPase.
History
DepositionNov 12, 2024-
Header (metadata) releaseMar 12, 2025-
Map releaseMar 12, 2025-
UpdateMar 12, 2025-
Current statusMar 12, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_62369.png

Downloads & links

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Map

FileDownload / File: emd_62369.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 384 pix.
= 318.72 Å		0.83 Å/pix.
x 384 pix.
= 318.72 Å		0.83 Å/pix.
x 384 pix.
= 318.72 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.09
Minimum - Maximum-0.19092816 - 0.6711576
Average (Standard dev.)0.0013787021 (±0.021167345)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 318.72 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_62369_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_62369_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human PNPase in open form

EntireName: Human PNPase in open form
Components
  • Complex: Human PNPase in open form
    • Protein or peptide: Human polynucleotide phosphorylase (hPNPase)

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Supramolecule #1: Human PNPase in open form

SupramoleculeName: Human PNPase in open form / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Human polynucleotide phosphorylase (hPNPase)

MacromoleculeName: Human polynucleotide phosphorylase (hPNPase) / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString: MGAVAVDLGN RKLEISSGKL ARFADGSAVV QSGDTAVMVT AVSKTKPSPS QFMPLVVDYR QKAAAAGRI PTNYLRREVG TSDKEILTSR IIDRSIRPLF PAGYFYDTQV LCNLLAVDGV N EPDVLAIN GASVALSLSD IPWNGPVGAV RIGIIDGEYV VNPTRKEMSS ...String:
MGAVAVDLGN RKLEISSGKL ARFADGSAVV QSGDTAVMVT AVSKTKPSPS QFMPLVVDYR QKAAAAGRI PTNYLRREVG TSDKEILTSR IIDRSIRPLF PAGYFYDTQV LCNLLAVDGV N EPDVLAIN GASVALSLSD IPWNGPVGAV RIGIIDGEYV VNPTRKEMSS STLNLVVAGA PK SQIVMLE ASAENILQQD FCHAIKVGVK YTQQIIQGIQ QLVKETGVTK RTPQKLFTPS PEI VKYTHK LAMERLYAVF TDYEHDKVSR DEAVNKIRLD TEEQLKEKFP EADPYEIIES FNVV AKEVF RSIVLNEYKR CDGRDLTSLR NVSCEVDMFK TLHGSALFQR GQTQVLCTVT FDSLE SGIK SDQVITAING IKDKNFMLHY EFPPYATNEI GKVTGLNRRE LGHGALAEKA LYPVIP RDF PFTIRVTSEV LESNGSSSMA SACGGSLALM DSGVPISSAV AGVAIGLVTK TDPEKGE IE DYRLLTDILG IEDYNGDMDF KIAGTNKGIT ALQADIKLPG IPIKIVMEAI QQASVAKK E ILQIMNKTIS KPRASRKENG PVVETVQVPL SKRAKFVGPG GYNLKKLQAE TGVTISQVD EETFSVFAPT PSAMHEARDF ITEICKDDQE QQLEFGAVYT ATITEIRDTG VMVKLYPNMT AVLLHNTQL DQRKIKHPTA LGLEVGQEIQ VKYFGRDPAD GRMRLSRKVL QSPATTVVRT L NDRSSIVM GEPISQSSSN SQAAALEHHH HHH

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.72 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 1357000
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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