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- EMDB-28275: Homocitrate-deficient nitrogenase MoFe-protein from A. vinelandii... -
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Open data
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Basic information
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Title | Homocitrate-deficient nitrogenase MoFe-protein from A. vinelandii nifV knockout in complex with NafT | ||||||||||||
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![]() | nitrogenase / nitrogen fixation / reductase / MoFe / OXIDOREDUCTASE | ||||||||||||
Function / homology | ![]() molybdenum-iron nitrogenase complex / nitrogenase / : / nitrogenase activity / nitrogen fixation / iron-sulfur cluster binding / ATP binding / metal ion binding Similarity search - Function | ||||||||||||
Biological species | ![]() | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.71 Å | ||||||||||||
![]() | Warmack RA / Maggiolo AO / Rees DC | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural consequences of turnover-induced homocitrate loss in nitrogenase. Authors: Rebeccah A Warmack / Ailiena O Maggiolo / Andres Orta / Belinda B Wenke / James B Howard / Douglas C Rees / ![]() Abstract: Nitrogenase catalyzes the ATP-dependent reduction of dinitrogen to ammonia during the process of biological nitrogen fixation that is essential for sustaining life. The active site FeMo-cofactor ...Nitrogenase catalyzes the ATP-dependent reduction of dinitrogen to ammonia during the process of biological nitrogen fixation that is essential for sustaining life. The active site FeMo-cofactor contains a [7Fe:1Mo:9S:1C] metallocluster coordinated with an R-homocitrate (HCA) molecule. Here, we establish through single particle cryoEM and chemical analysis of two forms of the Azotobacter vinelandii MoFe-protein - a high pH turnover inactivated species and a ∆NifV variant that cannot synthesize HCA - that loss of HCA is coupled to α-subunit domain and FeMo-cofactor disordering, and formation of a histidine coordination site. We further find a population of the ∆NifV variant complexed to an endogenous protein identified through structural and proteomic approaches as the uncharacterized protein NafT. Recognition by endogenous NafT demonstrates the physiological relevance of the HCA-compromised form, perhaps for cofactor insertion or repair. Our results point towards a dynamic active site in which HCA plays a role in enabling nitrogenase catalysis by facilitating activation of the FeMo-cofactor from a relatively stable form to a state capable of reducing dinitrogen under ambient conditions. | ||||||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 372 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 21 KB 21 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 19.1 KB | Display | ![]() |
Images | ![]() | 59.9 KB | ||
Filedesc metadata | ![]() | 6.3 KB | ||
Others | ![]() ![]() ![]() | 370.7 MB 675.7 MB 675.7 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8enoMC ![]() 8crsC ![]() 8dbxC ![]() 8enlC ![]() 8enmC ![]() 8ennC C: citing same article ( M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Voxel size | X=Y=Z: 0.65 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: Blurred version of the main map.
File | emd_28275_additional_1.map | ||||||||||||
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Annotation | Blurred version of the main map. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_28275_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_28275_half_map_2.map | ||||||||||||
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Density Histograms |
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Sample components
-Entire : Heterotetrameric nitrogenase MoFe-protein in complex with NafT is...
Entire | Name: Heterotetrameric nitrogenase MoFe-protein in complex with NafT isolated from nifV mutant strain |
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Components |
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-Supramolecule #1: Heterotetrameric nitrogenase MoFe-protein in complex with NafT is...
Supramolecule | Name: Heterotetrameric nitrogenase MoFe-protein in complex with NafT isolated from nifV mutant strain type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: ![]() |
-Macromolecule #1: Nitrogenase molybdenum-iron protein alpha chain
Macromolecule | Name: Nitrogenase molybdenum-iron protein alpha chain / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: nitrogenase |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 54.000559 KDa |
Sequence | String: MSREEVESLI QEVLEVYPEK ARKDRNKHLA VNDPAVTQSK KCIISNKKSQ PGLMTIRGCA YAGSKGVVWG PIKDMIHISH GPVGCGQYS RAGRRNYYIG TTGVNAFVTM NFTSDFQEKD IVFGGDKKLA KLIDEVETLF PLNKGISVQS ECPIGLIGDD I ESVSKVKG ...String: MSREEVESLI QEVLEVYPEK ARKDRNKHLA VNDPAVTQSK KCIISNKKSQ PGLMTIRGCA YAGSKGVVWG PIKDMIHISH GPVGCGQYS RAGRRNYYIG TTGVNAFVTM NFTSDFQEKD IVFGGDKKLA KLIDEVETLF PLNKGISVQS ECPIGLIGDD I ESVSKVKG AELSKTIVPV RCEGFRGVSQ SLGHHIANDA VRDWVLGKRD EDTTFASTPY DVAIIGDYNI GGDAWSSRIL LE EMGLRCV AQWSGDGSIS EIELTPKVKL NLVHCYRSMN YISRHMEEKY GIPWMEYNFF GPTKTIESLR AIAAKFDESI QKK CEEVIA KYKPEWEAVV AKYRPRLEGK RVMLYIGGLR PRHVIGAYED LGMEVVGTGY EFAHNDDYDR TMKEMGDSTL LYDD VTGYE FEEFVKRIKP DLIGSGIKEK FIFQKMGIPF REMHSWDYSG PYHGFDGFAI FARDMDMTLN NPCWKKLQAP WE UniProtKB: Nitrogenase molybdenum-iron protein alpha chain |
-Macromolecule #2: Nitrogenase molybdenum-iron protein beta chain
Macromolecule | Name: Nitrogenase molybdenum-iron protein beta chain / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO / EC number: nitrogenase |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 59.404684 KDa |
Sequence | String: SQQVDKIKAS YPLFLDQDYK DMLAKKRDGF EEKYPQDKID EVFQWTTTKE YQELNFQREA LTVNPAKACQ PLGAVLCALG FEKTMPYVH GSQGCVAYFR SYFNRHFREP VSCVSDSMTE DAAVFGGQQN MKDGLQNCKA TYKPDMIAVS TTCMAEVIGD D LNAFINNS ...String: SQQVDKIKAS YPLFLDQDYK DMLAKKRDGF EEKYPQDKID EVFQWTTTKE YQELNFQREA LTVNPAKACQ PLGAVLCALG FEKTMPYVH GSQGCVAYFR SYFNRHFREP VSCVSDSMTE DAAVFGGQQN MKDGLQNCKA TYKPDMIAVS TTCMAEVIGD D LNAFINNS KKEGFIPDEF PVPFAHTPSF VGSHVTGWDN MFEGIARYFT LKSMDDKVVG SNKKINIVPG FETYLGNFRV IK RMLSEMG VGYSLLSDPE EVLDTPADGQ FRMYAGGTTQ EEMKDAPNAL NTVLLQPWHL EKTKKFVEGT WKHEVPKLNI PMG LDWTDE FLMKVSEISG QPIPASLTKE RGRLVDMMTD SHTWLHGKRF ALWGDPDFVM GLVKFLLELG CEPVHILCHN GNKR WKKAV DAILAASPYG KNATVYIGKD LWHLRSLVFT DKPDFMIGNS YGKFIQRDTL HKGKEFEVPL IRIGFPIFDR HHLHR STTL GYEGAMQILT TLVNSILERL DEETRGMQAT DYNHDLVR UniProtKB: Nitrogenase molybdenum-iron protein beta chain |
-Macromolecule #3: nitrogenase-associated factor T
Macromolecule | Name: nitrogenase-associated factor T / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 14.847405 KDa |
Sequence | String: MSWRILLCHK HPVSARLRFL IPTGGGVVLP QTLPRLAVIA EDQEAPVQCH PASALRALQE TMALGWQLEL IGEFRLNMEV PGQIMPIYL AALAGHELPP PPEGTRWIEL TQSIGMPWLD RELLRRVYEE LIG UniProtKB: Uncharacterized protein |
-Macromolecule #4: iron-sulfur-molybdenum cluster with interstitial carbon
Macromolecule | Name: iron-sulfur-molybdenum cluster with interstitial carbon type: ligand / ID: 4 / Number of copies: 2 / Formula: ICS |
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Molecular weight | Theoretical: 787.451 Da |
Chemical component information | ![]() ChemComp-ICE: |
-Macromolecule #5: CITRIC ACID
Macromolecule | Name: CITRIC ACID / type: ligand / ID: 5 / Number of copies: 1 / Formula: CIT |
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Molecular weight | Theoretical: 192.124 Da |
Chemical component information | ![]() ChemComp-CIT: |
-Macromolecule #6: FE(8)-S(7) CLUSTER
Macromolecule | Name: FE(8)-S(7) CLUSTER / type: ligand / ID: 6 / Number of copies: 2 / Formula: CLF |
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Molecular weight | Theoretical: 671.215 Da |
Chemical component information | ![]() ChemComp-CLF: |
-Macromolecule #7: CHAPSO
Macromolecule | Name: CHAPSO / type: ligand / ID: 7 / Number of copies: 2 / Formula: 1N7 |
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Molecular weight | Theoretical: 631.884 Da |
Chemical component information | ![]() ChemComp-1N7: |
-Macromolecule #8: FE (III) ION
Macromolecule | Name: FE (III) ION / type: ligand / ID: 8 / Number of copies: 2 / Formula: FE |
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Molecular weight | Theoretical: 55.845 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 2.41 mg/mL |
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Buffer | pH: 7.8 |
Vitrification | Cryogen name: ETHANE-PROPANE |
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Electron microscopy
Microscope | TFS KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: -3.0 µm / Nominal defocus min: -0.8 µm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |