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- PDB-9cl6: Ammonia monooxygenase in native membranes -

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Basic information

Entry
Database: PDB / ID: 9cl6
TitleAmmonia monooxygenase in native membranes
Components(Ammonia monooxygenase ...) x 4
KeywordsOXIDOREDUCTASE / membrane protein / metalloenzyme / monooxygenase
Function / homology
Function and homology information


ammonia monooxygenase / ammonia monooxygenase activity / monooxygenase activity / metal ion binding / membrane / plasma membrane / cytoplasm
Similarity search - Function
Ammonia monooxygenase/particulate methane monooxygenase, subunit A / Ammonia monooxygenase/particulate methane monooxygenase, subunit C / Ammonia/methane monooxygenase, subunit B, hairpin domain superfamily / Ammonia/methane monooxygenase, subunit B, C-terminal / Ammonia monooxygenase/particulate methane monooxygenase, subunit C domain superfamily / Ammonia/particulate methane monooxygenase, subunit A superfamily / Ammonia monooxygenase / Ammonia monooxygenase/methane monooxygenase, subunit C / Ammonia monooxygenase/particulate methane monooxygenase, subunit B / Ammonia/methane monooxygenase, subunitB, N-terminal / Monooxygenase subunit B protein
Similarity search - Domain/homology
COPPER (II) ION / Ammonia monooxygenase alpha subunit / Ammonia monooxygenase beta subunit / Ammonia monooxygenase subunit C
Similarity search - Component
Biological speciesNitrosomonas europaea ATCC 19718 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsTucci, F.J. / Rosenzweig, A.C.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM118035 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)T32GM105538 United States
National Institutes of Health/National Institute of Environmental Health Sciences (NIH/NIEHS)F31ES034283 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Structures of methane and ammonia monooxygenases in native membranes.
Authors: Frank J Tucci / Amy C Rosenzweig /
Abstract: Methane- and ammonia-oxidizing bacteria play key roles in the global carbon and nitrogen cycles, respectively. These bacteria use homologous copper membrane monooxygenases to accomplish the defining ...Methane- and ammonia-oxidizing bacteria play key roles in the global carbon and nitrogen cycles, respectively. These bacteria use homologous copper membrane monooxygenases to accomplish the defining chemical transformations of their metabolisms: the oxidations of methane to methanol by particulate methane monooxygenase (pMMO) and ammonia to hydroxylamine by ammonia monooxygenase (AMO), enzymes of prime interest for applications in mitigating climate change. However, investigations of these enzymes have been hindered by the need for disruptive detergent solubilization prior to structure determination, confounding studies of pMMO and precluding studies of AMO. Here, we overcome these challenges by using cryoEM to visualize pMMO and AMO directly in their native membrane arrays at 2.4 to 2.8 Å resolution. These structures reveal details of the copper centers, numerous bound lipids, and previously unobserved components, including identifiable and distinct supernumerary helices interacting with pMMO and AMO, suggesting a widespread role for these helices in copper membrane monooxygenases. Comparisons between these structures, their metallocofactors, and their unexpected protein-protein interactions highlight features that may govern activity or the formation of higher-order arrays in native membranes. The ability to obtain molecular insights within the native membrane will enable further understanding of these environmentally important enzymes.
History
DepositionJul 10, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 8, 2025Provider: repository / Type: Initial release
Revision 1.1Jan 15, 2025Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: Ammonia monooxygenase subunit C
A: Ammonia monooxygenase beta subunit
B: Ammonia monooxygenase alpha subunit
Da: ammonia monooxygenase supernumerary helix
Db: ammonia monooxygenase supernumerary helix
Dc: ammonia monooxygenase supernumerary helix
I: Ammonia monooxygenase alpha subunit
F: Ammonia monooxygenase alpha subunit
J: Ammonia monooxygenase subunit C
G: Ammonia monooxygenase subunit C
H: Ammonia monooxygenase beta subunit
E: Ammonia monooxygenase beta subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)327,08718
Polymers326,70612
Non-polymers3816
Water14,808822
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Ammonia monooxygenase ... , 4 types, 12 molecules CJGAHEBIFDaDbDc

#1: Protein Ammonia monooxygenase subunit C


Mass: 29956.896 Da / Num. of mol.: 3 / Source method: isolated from a natural source
Source: (natural) Nitrosomonas europaea ATCC 19718 (bacteria)
References: UniProt: Q82T63
#2: Protein Ammonia monooxygenase beta subunit / AMO / Heterotrimeric Cu-heme enzyme


Mass: 43037.168 Da / Num. of mol.: 3 / Source method: isolated from a natural source
Source: (natural) Nitrosomonas europaea ATCC 19718 (bacteria)
References: UniProt: Q04508, ammonia monooxygenase
#3: Protein Ammonia monooxygenase alpha subunit / AMO / Acetylene-binding polypeptide / Heterotrimeric Cu-heme enzyme


Mass: 31800.271 Da / Num. of mol.: 3 / Source method: isolated from a natural source
Source: (natural) Nitrosomonas europaea ATCC 19718 (bacteria)
References: UniProt: Q04507, ammonia monooxygenase
#4: Protein/peptide ammonia monooxygenase supernumerary helix


Mass: 4107.730 Da / Num. of mol.: 3 / Source method: obtained synthetically
Source: (synth.) Nitrosomonas europaea ATCC 19718 (bacteria)

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Non-polymers , 2 types, 828 molecules

#5: Chemical
ChemComp-CU / COPPER (II) ION


Mass: 63.546 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Cu / Feature type: SUBJECT OF INVESTIGATION
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 822 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: 2D ARRAY / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: ammonia monooxygenase in native membranes / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1-#4 / Source: NATURAL
Source (natural)Organism: Nitrosomonas europaea ATCC 19718 (bacteria)
Buffer solutionpH: 7.2
SpecimenConc.: 4 mg/ml / Embedding applied: YES / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3
EM embeddingMaterial: vitreous ice
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.21_5207: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 79436 / Symmetry type: POINT

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