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- EMDB-37639: Cryo-EM structure of ACE2-SIT1 complex with tiagabine -

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Entry
Database: EMDB / ID: EMD-37639
TitleCryo-EM structure of ACE2-SIT1 complex with tiagabine
Map data
Sample
  • Complex: ACE2-SIT1 compelx with tiagabine
    • Protein or peptide: Sodium- and chloride-dependent transporter XTRP3
    • Protein or peptide: Angiotensin-converting enzyme 2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: Tiagabine
KeywordsTransporter / inhibitors / TRANSPORT PROTEIN / TRANSPORT PROTEIN-HYDROLASE complex
Function / homology
Function and homology information


proline:sodium symporter activity / L-isoleucine transmembrane transporter activity / solute:sodium symporter activity / L-isoleucine import across plasma membrane / L-proline import across plasma membrane / Variant SLC6A20 contributes towards hyperglycinuria (HG) and iminoglycinuria (IG) / Variant SLC6A20 contributes towards hyperglycinuria (HG) and iminoglycinuria (IG) / amino-acid betaine transport / proline import across plasma membrane / L-proline transmembrane transporter activity ...proline:sodium symporter activity / L-isoleucine transmembrane transporter activity / solute:sodium symporter activity / L-isoleucine import across plasma membrane / L-proline import across plasma membrane / Variant SLC6A20 contributes towards hyperglycinuria (HG) and iminoglycinuria (IG) / Variant SLC6A20 contributes towards hyperglycinuria (HG) and iminoglycinuria (IG) / amino-acid betaine transport / proline import across plasma membrane / L-proline transmembrane transporter activity / glycine import across plasma membrane / amino-acid betaine transmembrane transporter activity / glycine transport / amino acid import across plasma membrane / proline transport / amino acid transmembrane transporter activity / Amino acid transport across the plasma membrane / neutral L-amino acid transmembrane transporter activity / Na+/Cl- dependent neurotransmitter transporters / amino acid transport / positive regulation of amino acid transport / angiotensin-converting enzyme 2 / positive regulation of L-proline import across plasma membrane / Hydrolases; Acting on peptide bonds (peptidases); Metallocarboxypeptidases / angiotensin-mediated drinking behavior / regulation of systemic arterial blood pressure by renin-angiotensin / tryptophan transport / positive regulation of gap junction assembly / transport across blood-brain barrier / regulation of cardiac conduction / regulation of vasoconstriction / peptidyl-dipeptidase activity / maternal process involved in female pregnancy / sodium ion transmembrane transport / angiotensin maturation / Metabolism of Angiotensinogen to Angiotensins / negative regulation of signaling receptor activity / carboxypeptidase activity / Attachment and Entry / positive regulation of cardiac muscle contraction / viral life cycle / regulation of cytokine production / blood vessel diameter maintenance / negative regulation of smooth muscle cell proliferation / brush border membrane / regulation of transmembrane transporter activity / cilium / negative regulation of ERK1 and ERK2 cascade / metallopeptidase activity / positive regulation of reactive oxygen species metabolic process / endocytic vesicle membrane / virus receptor activity / regulation of cell population proliferation / regulation of inflammatory response / endopeptidase activity / Potential therapeutics for SARS / Induction of Cell-Cell Fusion / entry receptor-mediated virion attachment to host cell / receptor-mediated endocytosis of virus by host cell / membrane fusion / Attachment and Entry / receptor-mediated virion attachment to host cell / symbiont entry into host cell / membrane raft / apical plasma membrane / endoplasmic reticulum lumen / cell surface / extracellular space / extracellular exosome / zinc ion binding / extracellular region / identical protein binding / membrane / plasma membrane
Similarity search - Function
Neutral amino acid SLC6 transporter / Sodium:neurotransmitter symporter family signature 2. / Sodium:neurotransmitter symporter family signature 1. / Sodium:neurotransmitter symporter / Sodium:neurotransmitter symporter superfamily / Sodium:neurotransmitter symporter family / Sodium:neurotransmitter symporter family profile. / Collectrin domain / Renal amino acid transporter / Collectrin-like domain profile. ...Neutral amino acid SLC6 transporter / Sodium:neurotransmitter symporter family signature 2. / Sodium:neurotransmitter symporter family signature 1. / Sodium:neurotransmitter symporter / Sodium:neurotransmitter symporter superfamily / Sodium:neurotransmitter symporter family / Sodium:neurotransmitter symporter family profile. / Collectrin domain / Renal amino acid transporter / Collectrin-like domain profile. / Peptidase M2, peptidyl-dipeptidase A / Angiotensin-converting enzyme / Peptidase family M2 domain profile. / Neutral zinc metallopeptidases, zinc-binding region signature.
Similarity search - Domain/homology
Angiotensin-converting enzyme 2 / Sodium- and chloride-dependent transporter XTRP3
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.34 Å
AuthorsYan R / Hu Z / Dai L / Zhang T
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32371267 China
CitationJournal: J Biol Chem / Year: 2024
Title: Cryo-EM structure of ACE2-SIT1 in complex with tiagabine.
Authors: Angelika Bröer / Ziwei Hu / Jędrzej Kukułowicz / Aditya Yadav / Ting Zhang / Lu Dai / Marek Bajda / Renhong Yan / Stefan Bröer /
Abstract: The pharmacology of amino acid transporters in the SLC6 family is poorly developed compared to that of the neurotransmitter transporters. To identify new inhibitors of the proline transporter SIT1 ...The pharmacology of amino acid transporters in the SLC6 family is poorly developed compared to that of the neurotransmitter transporters. To identify new inhibitors of the proline transporter SIT1 (SLC6A20), its expression in Xenopus laevis oocytes was optimized. Trafficking of SIT1 was augmented by co-expression of angiotensin-converting enzyme 2 (ACE2) in oocytes but there was no strict requirement for co-expression of ACE2. A pharmacophore-guided screen identified tiagabine as a potent non-competitive inhibitor of SIT1. To understand its binding mode, we determined the cryo-electron microscopy (cryo-EM) structure of ACE2-SIT1 bound with tiagabine. The inhibitor binds close to the orthosteric proline binding site, but due to its size extends into the cytosolic vestibule. This causes the transporter to adopt an inward-open conformation, in which the intracellular gate is blocked. This study provides the first structural insight into inhibition of SIT1 and generates tools for a better understanding of the ACE2-SIT1 complex. These findings may have significance for SARS-CoV-2 binding to its receptor ACE2 in human lung alveolar cells where SIT1 and ACE2 are functionally expressed.
History
DepositionOct 2, 2023-
Header (metadata) releaseSep 25, 2024-
Map releaseSep 25, 2024-
UpdateSep 25, 2024-
Current statusSep 25, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_37639.png

Downloads & links

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Map

FileDownload / File: emd_37639.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 256 pix.
= 280.32 Å		1.1 Å/pix.
x 256 pix.
= 280.32 Å		1.1 Å/pix.
x 256 pix.
= 280.32 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.095 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.247
Minimum - Maximum-0.26774445 - 0.9055063
Average (Standard dev.)0.009450303 (±0.051834557)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 280.32 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #1

Fileemd_37639_additional_1.map
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Half map: #2

Fileemd_37639_half_map_1.map
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Half map: #1

Fileemd_37639_half_map_2.map
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Sample components

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Entire : ACE2-SIT1 compelx with tiagabine

EntireName: ACE2-SIT1 compelx with tiagabine
Components
  • Complex: ACE2-SIT1 compelx with tiagabine
    • Protein or peptide: Sodium- and chloride-dependent transporter XTRP3
    • Protein or peptide: Angiotensin-converting enzyme 2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: Tiagabine

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Supramolecule #1: ACE2-SIT1 compelx with tiagabine

SupramoleculeName: ACE2-SIT1 compelx with tiagabine / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Sodium- and chloride-dependent transporter XTRP3

MacromoleculeName: Sodium- and chloride-dependent transporter XTRP3 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 68.010492 KDa
Recombinant expressionOrganism: Eukalyptorhynchia (invertebrata)
SequenceString: DYKDDDDKSG PDEVDASGRM EKARPLWANS LQFVFACTSY AVGLGNVWRF PYLCQMYGGG SFLVPYIIML IVEGMPLLYL ELAVGQRMR QGSIGAWRTI SPYLSGVGVA SVVVSFFLSM YYNVINAWAF WYLFHSFQDP LPWSVCPLNG NHTGYDEECE K ASSTQYFW ...String:
DYKDDDDKSG PDEVDASGRM EKARPLWANS LQFVFACTSY AVGLGNVWRF PYLCQMYGGG SFLVPYIIML IVEGMPLLYL ELAVGQRMR QGSIGAWRTI SPYLSGVGVA SVVVSFFLSM YYNVINAWAF WYLFHSFQDP LPWSVCPLNG NHTGYDEECE K ASSTQYFW YRKTLNISPS LQENGGVQWE PALCLLLAWL VVYLCILRGT ESTGKVVYFT ASLPYCVLII YLIRGLTLHG AT NGLMYMF TPKIEQLANP KAWINAATQI FFSLGLGFGS LIAFASYNEP SNNCQKHAII VSLINSFTSI FASIVTFSIY GFK ATFNYE NCLKKVSLLL TNTFDLEDGF LTASNLEQVK GYLASAYPSK YSEMFPQIKN CSLESELDTA VQGTGLAFIV YTEA IKNME VSQLWSVLYF FMLLMLGIGS MLGNTAAILT PLTDSKIISS HLPKEAISGL VCLVNCAIGM VFTMEAGNYW FDIFN DYAA TLSLLLIVLV ETIAVCYVYG LRRFESDLKA MTGRAVSWYW KVMWAGVSPL LIVSLFVFYL SDYILTGTLK YQAWDA SQG QLVTKDYPAY ALAVIGLLVA SSTMCIPLAA LGTFVQRRLK RGDADPVA

UniProtKB: Sodium- and chloride-dependent transporter XTRP3

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Macromolecule #2: Angiotensin-converting enzyme 2

MacromoleculeName: Angiotensin-converting enzyme 2 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO / EC number: angiotensin-converting enzyme 2
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 94.16643 KDa
Recombinant expressionOrganism: Eukalyptorhynchia (invertebrata)
SequenceString: MSSSSWLLLS LVAVTAAQSS IEEQAKTFLD KFNHEAEDLF YQSSLASWNY NTNITEENVQ NMNNAGDKWS AFLKEQSTLA QMYPLQEIQ NLTVKLQLQA LQQNGSSVLS EDKSKRLNTI LNTMSTIYST GKVCNPDNPQ ECLLLEPGLN EIMANSLDYN E RLWAWESW ...String:
MSSSSWLLLS LVAVTAAQSS IEEQAKTFLD KFNHEAEDLF YQSSLASWNY NTNITEENVQ NMNNAGDKWS AFLKEQSTLA QMYPLQEIQ NLTVKLQLQA LQQNGSSVLS EDKSKRLNTI LNTMSTIYST GKVCNPDNPQ ECLLLEPGLN EIMANSLDYN E RLWAWESW RSEVGKQLRP LYEEYVVLKN EMARANHYED YGDYWRGDYE VNGVDGYDYS RGQLIEDVEH TFEEIKPLYE HL HAYVRAK LMNAYPSYIS PIGCLPAHLL GDMWGRFWTN LYSLTVPFGQ KPNIDVTDAM VDQAWDAQRI FKEAEKFFVS VGL PNMTQG FWENSMLTDP GNVQKAVCHP TAWDLGKGDF RILMCTKVTM DDFLTAHHEM GHIQYDMAYA AQPFLLRNGA NEGF HEAVG EIMSLSAATP KHLKSIGLLS PDFQEDNETE INFLLKQALT IVGTLPFTYM LEKWRWMVFK GEIPKDQWMK KWWEM KREI VGVVEPVPHD ETYCDPASLF HVSNDYSFIR YYTRTLYQFQ FQEALCQAAK HEGPLHKCDI SNSTEAGQKL FNMLRL GKS EPWTLALENV VGAKNMNVRP LLNYFEPLFT WLKDQNKNSF VGWSTDWSPY ADQSIKVRIS LKSALGDKAY EWNDNEM YL FRSSVAYAMR QYFLKVKNQM ILFGEEDVRV ANLKPRISFN FFVTAPKNVS DIIPRTEVEK AIRMSRSRIN DAFRLNDN S LEFLGIQPTL GPPNQPPVSI WLIVFGVVMG VIVVGIVILI FTGIRDRKKK NKARSGENPY ASIDISKGEN NPGFQNTDD VQTSFLEHHH HHHHHHH

UniProtKB: Angiotensin-converting enzyme 2

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 4 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #5: Tiagabine

MacromoleculeName: Tiagabine / type: ligand / ID: 5 / Number of copies: 2 / Formula: TGI
Molecular weightTheoretical: 375.548 Da
Chemical component information

ChemComp-TGI:
Tiagabine / medication*YM

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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: 7.5
VitrificationCryogen name: NITROGEN

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number real images: 1848 / Average exposure time: 1.873 sec. / Average electron dose: 1.5625 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 1.4000000000000001 µm / Nominal magnification: 81000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

33652

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.34 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 193320
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: COMMON LINE

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