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- PDB-8sr1: particulate methane monooxygenase crosslinked with 4,4,4-trifluor... -

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Basic information

Entry
Database: PDB / ID: 8sr1
Titleparticulate methane monooxygenase crosslinked with 4,4,4-trifluorobutanol bound
Components
  • (Particulate methane monooxygenase ...) x 2
  • Ammonia monooxygenase/methane monooxygenase, subunit C family protein
KeywordsOXIDOREDUCTASE / Metalloenzyme / Inhibitor / Membrane Protein / Nanodiscs
Function / homology
Function and homology information


methane monooxygenase (particulate) / methane monooxygenase (soluble) / methane monooxygenase [NAD(P)H] activity / monooxygenase activity / membrane / metal ion binding
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 / DECANE / 1,2-dihexanoyl-sn-glycero-3-phosphocholine / 1,2-DIDECANOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / DIUNDECYL PHOSPHATIDYL CHOLINE / 4,4,4-trifluorobutan-1-ol / Particulate methane monooxygenase alpha subunit / Ammonia monooxygenase/methane monooxygenase, subunit C family protein / Particulate methane monooxygenase beta subunit
Similarity search - Component
Biological speciesMethylococcus capsulatus str. Bath (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.18 Å
AuthorsTucci, F.J. / Rosenzweig, A.C.
Funding support United States, 6items
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)R01GM111097 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
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)T32GM008382 United States
National Science Foundation (NSF, United States)MCB-1908587 United States
CitationJournal: Nat Catal / Year: 2023
Title: Product analog binding identifies the copper active site of particulate methane monooxygenase.
Authors: Frank J Tucci / Richard J Jodts / Brian M Hoffman / Amy C Rosenzweig /
Abstract: Nature's primary methane-oxidizing enzyme, the membrane-bound particulate methane monooxygenase (pMMO), catalyzes the oxidation of methane to methanol. pMMO activity requires copper, and decades of ...Nature's primary methane-oxidizing enzyme, the membrane-bound particulate methane monooxygenase (pMMO), catalyzes the oxidation of methane to methanol. pMMO activity requires copper, and decades of structural and spectroscopic studies have sought to identify the active site among three candidates: the Cu, Cu, and Cu sites. Challenges associated with the isolation of active pMMO have hindered progress toward locating its catalytic center. However, reconstituting pMMO into native lipid nanodiscs stabilizes its structure and recovers its activity. Here, these active samples were incubated with 2,2,2,-trifluoroethanol (TFE), a product analog that serves as a readily visualized active-site probe. Interactions of TFE with the Cu site were observed by both pulsed ENDOR spectroscopy and cryoEM, implicating Cu and the surrounding hydrophobic pocket as the likely site of methane oxidation. Use of these orthogonal techniques on parallel samples is a powerful approach that can circumvent difficulties in interpreting metalloenzyme cryoEM maps.
History
DepositionMay 5, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 15, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 24, 2024Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Particulate methane monooxygenase alpha subunit
B: Particulate methane monooxygenase beta subunit
C: Ammonia monooxygenase/methane monooxygenase, subunit C family protein
E: Particulate methane monooxygenase alpha subunit
F: Particulate methane monooxygenase beta subunit
I: Particulate methane monooxygenase alpha subunit
J: Particulate methane monooxygenase beta subunit
G: Ammonia monooxygenase/methane monooxygenase, subunit C family protein
K: Ammonia monooxygenase/methane monooxygenase, subunit C family protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)319,33475
Polymers295,0789
Non-polymers24,25666
Water6,323351
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Particulate methane monooxygenase ... , 2 types, 6 molecules AEIBFJ

#1: Protein Particulate methane monooxygenase alpha subunit


Mass: 42832.887 Da / Num. of mol.: 3 / Source method: isolated from a natural source
Source: (natural) Methylococcus capsulatus str. Bath (bacteria)
References: UniProt: G1UBD1
#2: Protein Particulate methane monooxygenase beta subunit


Mass: 27869.459 Da / Num. of mol.: 3 / Source method: isolated from a natural source
Source: (natural) Methylococcus capsulatus str. Bath (bacteria)
References: UniProt: Q607G3

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Protein , 1 types, 3 molecules CGK

#3: Protein Ammonia monooxygenase/methane monooxygenase, subunit C family protein


Mass: 27656.840 Da / Num. of mol.: 3 / Source method: isolated from a natural source
Source: (natural) Methylococcus capsulatus str. Bath (bacteria)
References: UniProt: Q603F1

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Non-polymers , 7 types, 417 molecules

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


Mass: 63.546 Da / Num. of mol.: 9 / Source method: obtained synthetically / Formula: Cu
#5: Chemical
ChemComp-D10 / DECANE


Mass: 142.282 Da / Num. of mol.: 18 / Source method: obtained synthetically / Formula: C10H22
#6: Chemical
ChemComp-PLC / DIUNDECYL PHOSPHATIDYL CHOLINE


Mass: 622.834 Da / Num. of mol.: 18 / Source method: obtained synthetically / Formula: C32H65NO8P / Comment: phospholipid*YM
#7: Chemical
ChemComp-P1O / 1,2-DIDECANOYL-SN-GLYCERO-3-PHOSPHOCHOLINE


Mass: 566.728 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C28H57NO8P / Comment: DDPC, phospholipid*YM
#8: Chemical
ChemComp-HXG / 1,2-dihexanoyl-sn-glycero-3-phosphocholine / (4R,7R)-7-(hexanoyloxy)-4-hydroxy-N,N,N-trimethyl-10-oxo-3,5,9-trioxa-4-phosphapentadecan-1-aminium 4-oxide


Mass: 454.515 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C20H41NO8P
#9: Chemical ChemComp-WIY / 4,4,4-trifluorobutan-1-ol


Mass: 128.093 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C4H7F3O / Feature type: SUBJECT OF INVESTIGATION
#10: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 351 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY

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Experimental details

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Experiment

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

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Sample preparation

ComponentName: particulate methane monooxygenase / Type: COMPLEX / Entity ID: #1-#3 / Source: NATURAL
Molecular weightValue: 337.11 kDa/nm / Experimental value: NO
Source (natural)Organism: Methylococcus capsulatus str. Bath (bacteria)
Buffer solutionpH: 7.3
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: OTHER / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: OTHER / Nominal magnification: 81000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 53.55 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.18 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 890148 / Symmetry type: POINT

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