[English] 日本語
Yorodumi
- EMDB-25163: HtrA1S328A:Fab15H6.v4 complex -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-25163
TitleHtrA1S328A:Fab15H6.v4 complex
Map dataMain Map without enforced symmetry (C1). Used for structure determination
Sample
  • Complex: HtrA1PD/SA bound by Fab15H6.v4 at LoopA epitope
    • Protein or peptide: Serine protease HTRA1
    • Protein or peptide: Fab15H6.v4 Heavy Chain
    • Protein or peptide: Fab15H6.v4 Light Chain
Function / homology
Function and homology information


chorionic trophoblast cell differentiation / programmed cell death / growth factor binding / negative regulation of BMP signaling pathway / Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases / Degradation of the extracellular matrix / serine-type peptidase activity / placenta development / molecular function activator activity / negative regulation of transforming growth factor beta receptor signaling pathway ...chorionic trophoblast cell differentiation / programmed cell death / growth factor binding / negative regulation of BMP signaling pathway / Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases / Degradation of the extracellular matrix / serine-type peptidase activity / placenta development / molecular function activator activity / negative regulation of transforming growth factor beta receptor signaling pathway / collagen-containing extracellular matrix / positive regulation of apoptotic process / serine-type endopeptidase activity / proteolysis / extracellular space / extracellular exosome / extracellular region / identical protein binding / plasma membrane / cytosol
Similarity search - Function
Insulin-like growth factor binding protein / Insulin-like growth factor-binding protein, IGFBP / Insulin-like growth factor-binding protein (IGFBP) N-terminal domain profile. / Insulin growth factor-binding protein homologues / Kazal-type serine protease inhibitor domain / PDZ domain 6 / Kazal type serine protease inhibitors / PDZ domain / Peptidase S1C / Kazal domain superfamily ...Insulin-like growth factor binding protein / Insulin-like growth factor-binding protein, IGFBP / Insulin-like growth factor-binding protein (IGFBP) N-terminal domain profile. / Insulin growth factor-binding protein homologues / Kazal-type serine protease inhibitor domain / PDZ domain 6 / Kazal type serine protease inhibitors / PDZ domain / Peptidase S1C / Kazal domain superfamily / Trypsin-like peptidase domain / Kazal domain / Kazal domain profile. / Growth factor receptor cysteine-rich domain superfamily / PDZ domain profile. / Domain present in PSD-95, Dlg, and ZO-1/2. / PDZ domain / PDZ superfamily / Peptidase S1, PA clan
Similarity search - Domain/homology
Serine protease HTRA1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsGerhardy S / Green E / Estevez A / Arthur CP / Ultsch M / Rohou A / Kirchhofer D
Funding support United States, 1 items
OrganizationGrant numberCountry
Not funded United States
CitationJournal: Nat Commun / Year: 2022
Title: Allosteric inhibition of HTRA1 activity by a conformational lock mechanism to treat age-related macular degeneration.
Authors: Stefan Gerhardy / Mark Ultsch / Wanjian Tang / Evan Green / Jeffrey K Holden / Wei Li / Alberto Estevez / Chris Arthur / Irene Tom / Alexis Rohou / Daniel Kirchhofer /
Abstract: The trimeric serine protease HTRA1 is a genetic risk factor associated with geographic atrophy (GA), a currently untreatable form of age-related macular degeneration. Here, we describe the allosteric ...The trimeric serine protease HTRA1 is a genetic risk factor associated with geographic atrophy (GA), a currently untreatable form of age-related macular degeneration. Here, we describe the allosteric inhibition mechanism of HTRA1 by a clinical Fab fragment, currently being evaluated for GA treatment. Using cryo-EM, X-ray crystallography and biochemical assays we identify the exposed LoopA of HTRA1 as the sole Fab epitope, which is approximately 30 Å away from the active site. The cryo-EM structure of the HTRA1:Fab complex in combination with molecular dynamics simulations revealed that Fab binding to LoopA locks HTRA1 in a non-competent conformational state, incapable of supporting catalysis. Moreover, grafting the HTRA1-LoopA epitope onto HTRA2 and HTRA3 transferred the allosteric inhibition mechanism. This suggests a conserved conformational lock mechanism across the HTRA family and a critical role of LoopA for catalysis, which was supported by the reduced activity of HTRA1-3 upon LoopA deletion or perturbation. This study reveals the long-range inhibition mechanism of the clinical Fab and identifies an essential function of the exposed LoopA for activity of HTRA family proteases.
History
DepositionOct 18, 2021-
Header (metadata) releaseSep 7, 2022-
Map releaseSep 7, 2022-
UpdateSep 21, 2022-
Current statusSep 21, 2022Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_25163.png

Downloads & links

-
Map

FileDownload / File: emd_25163.map.gz / Format: CCP4 / Size: 8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMain Map without enforced symmetry (C1). Used for structure determination
Voxel sizeX=Y=Z: 1.2 Å
Density
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-1.0713027 - 1.3790567
Average (Standard dev.)0.00940577 (±0.07155112)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions128128128
Spacing128128128
CellA=B=C: 153.6 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Additional map: Half-maps 2/2 for additional map (C3)

Fileemd_25163_additional_1.map
AnnotationHalf-maps 2/2 for additional map (C3)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Additional map: Additional Map with enforced symmetry (C3).

Fileemd_25163_additional_2.map
AnnotationAdditional Map with enforced symmetry (C3).
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Additional map: Half-maps 1/2 for additional map (C3)

Fileemd_25163_additional_3.map
AnnotationHalf-maps 1/2 for additional map (C3)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Half-maps 2/2 for main map (C1)

Fileemd_25163_half_map_1.map
AnnotationHalf-maps 2/2 for main map (C1)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Half-maps 1/2 for main map (C1)

Fileemd_25163_half_map_2.map
AnnotationHalf-maps 1/2 for main map (C1)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : HtrA1PD/SA bound by Fab15H6.v4 at LoopA epitope

EntireName: HtrA1PD/SA bound by Fab15H6.v4 at LoopA epitope
Components
  • Complex: HtrA1PD/SA bound by Fab15H6.v4 at LoopA epitope
    • Protein or peptide: Serine protease HTRA1
    • Protein or peptide: Fab15H6.v4 Heavy Chain
    • Protein or peptide: Fab15H6.v4 Light Chain

-
Supramolecule #1: HtrA1PD/SA bound by Fab15H6.v4 at LoopA epitope

SupramoleculeName: HtrA1PD/SA bound by Fab15H6.v4 at LoopA epitope / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all / Details: clinical Fab fragment Fab15H6.v4
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)
Molecular weightTheoretical: 220 KDa

-
Macromolecule #1: Serine protease HTRA1

MacromoleculeName: Serine protease HTRA1 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
EC number: Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.714385 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHGEN LYFQGSDPNS LRHKYNFIAD VVEKIAPAVV HIELFRKLPF SKREVPVASG SGFIVSEDGL IVTNAHVVTN KHRVKVELK NGATYEAKIK DVDEKADIAL IKIDHQGKLP VLLLGRSSEL RPGEFVVAIG SPFSLQNTVT TGIVSTTQRG G KELGLRNS ...String:
MHHHHHHGEN LYFQGSDPNS LRHKYNFIAD VVEKIAPAVV HIELFRKLPF SKREVPVASG SGFIVSEDGL IVTNAHVVTN KHRVKVELK NGATYEAKIK DVDEKADIAL IKIDHQGKLP VLLLGRSSEL RPGEFVVAIG SPFSLQNTVT TGIVSTTQRG G KELGLRNS DMDYIQTDAI INYGNAGGPL VNLDGEVIGI NTLKVTAGIS FAIPSDKIKK FLTESHDRQA KGKAITK

-
Macromolecule #2: Fab15H6.v4 Heavy Chain

MacromoleculeName: Fab15H6.v4 Heavy Chain / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 26.768996 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKKNIAFLLA SMFVFSIATN AYAEVQLVQS GAEVKKPGAS VKVSCKASGY KFTDSEMHWV RQAPGQGLEW IGGVDPETEG AAYNQKFKG RATITRDTST STAYLELSSL RSEDTAVYYC TRGYDYDYAL DYWGQGTLVT VSSASTKGPS VFPLAPSSKS T SGGTAALG ...String:
MKKNIAFLLA SMFVFSIATN AYAEVQLVQS GAEVKKPGAS VKVSCKASGY KFTDSEMHWV RQAPGQGLEW IGGVDPETEG AAYNQKFKG RATITRDTST STAYLELSSL RSEDTAVYYC TRGYDYDYAL DYWGQGTLVT VSSASTKGPS VFPLAPSSKS T SGGTAALG CLVKDYFPEP VTVSWNSGAL TSGVHTFPAV LQSSGLYSLS SVVTVPSSSL GTQTYICNVN HKPSNTKVDK KV EPKSCDK THT

-
Macromolecule #3: Fab15H6.v4 Light Chain

MacromoleculeName: Fab15H6.v4 Light Chain / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.71773 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKKNIAFLLA SMFVFSIATN AYADIQMTQS PSSLSASVGD RVTITCRASS SVEFIHWYQQ KPGKAPKPLI SATSNLASGV PSRFSGSGS GTDFTLTISS LQPEDFATYY CQQWSSAPWT FGQGTKVEIK RTVAAPSVFI FPPSDEQLKS GTASVVCLLN N FYPREAKV ...String:
MKKNIAFLLA SMFVFSIATN AYADIQMTQS PSSLSASVGD RVTITCRASS SVEFIHWYQQ KPGKAPKPLI SATSNLASGV PSRFSGSGS GTDFTLTISS LQPEDFATYY CQQWSSAPWT FGQGTKVEIK RTVAAPSVFI FPPSDEQLKS GTASVVCLLN N FYPREAKV QWKVDNALQS GNSQESVTEQ DSKDSTYSLS STLTLSKADY EKHKVYACEV THQGLSSPVT KSFNRGEC

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration4 mg/mL
BufferpH: 7.2
Component:
ConcentrationFormulaName
200.0 mMNaClSodium chlorideSodium Chloride
50.0 mMTrisTris
0.25 PercentCHAPSCHAPS
GridModel: Quantifoil R0.6/1 / Material: GOLD / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.067 kPa
Details: SAM: Grids were incubated in 4 mM monothiolalkane(C11)PEG6-OH (11-mercaptoundecyl) hexaethylenglycol (SPT-0011P6, SensoPath Technologies, Inc., Bozeman, MT) for 24h and rinsed in EtOH before sample application
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: LEICA EM GP / Details: 3.5s blot time.
DetailsHtrA1PD/SA:Fab15H6.v4 complex

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 7336 / Average exposure time: 3.0 sec. / Average electron dose: 65.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Particle selectionNumber selected: 505669
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3tjn


Details: apo HtrA1
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM / Number images used: 219676

-
Atomic model buiding 1

Initial modelPDB ID:

3tjn


Chain - Chain ID: A
RefinementProtocol: RIGID BODY FIT
Output model

PDB-7sjo:
HtrA1S328A:Fab15H6.v4 complex

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more