National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R01 GM118396
United States
National Institutes of Health/National Cancer Institute (NIH/NCI)
CA197855
United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R01 GM047414
United States
Citation
Journal: J Cell Biol / Year: 2017 Title: The glycolytic enzyme phosphofructokinase-1 assembles into filaments. Authors: Bradley A Webb / Anne M Dosey / Torsten Wittmann / Justin M Kollman / Diane L Barber / Abstract: Despite abundant knowledge of the regulation and biochemistry of glycolytic enzymes, we have limited understanding on how they are spatially organized in the cell. Emerging evidence indicates that ...Despite abundant knowledge of the regulation and biochemistry of glycolytic enzymes, we have limited understanding on how they are spatially organized in the cell. Emerging evidence indicates that nonglycolytic metabolic enzymes regulating diverse pathways can assemble into polymers. We now show tetramer- and substrate-dependent filament assembly by phosphofructokinase-1 (PFK1), which is considered the "gatekeeper" of glycolysis because it catalyzes the step committing glucose to breakdown. Recombinant liver PFK1 (PFKL) isoform, but not platelet PFK1 (PFKP) or muscle PFK1 (PFKM) isoforms, assembles into filaments. Negative-stain electron micrographs reveal that filaments are apolar and made of stacked tetramers oriented with exposed catalytic sites positioned along the edge of the polymer. Electron micrographs and biochemical data with a PFKL/PFKP chimera indicate that the PFKL regulatory domain mediates filament assembly. Quantified live-cell imaging shows dynamic properties of localized PFKL puncta that are enriched at the plasma membrane. These findings reveal a new behavior of a key glycolytic enzyme with insights on spatial organization and isoform-specific glucose metabolism in cells.
History
Deposition
Dec 27, 2016
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Header (metadata) release
Jan 11, 2017
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Map release
Jan 24, 2018
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Update
Jan 29, 2020
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Current status
Jan 29, 2020
Processing site: RCSB / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Download / File: emd_8542.map.gz / Format: CCP4 / Size: 6.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation
wildtype phosphofructokinase liver isoform filaments assembled with ATP and fructose-6-phosphate
Voxel size
X=Y=Z: 4.14 Å
Density
Contour Level
By AUTHOR: 0.163 / Movie #1: 0.163
Minimum - Maximum
-0.2782783 - 0.5283528
Average (Standard dev.)
-0.00129166 (±0.059799593)
Symmetry
Space group: 1
Details
EMDB XML:
Map geometry
Axis order
X
Y
Z
Origin
-60
-60
-60
Dimensions
120
120
120
Spacing
120
120
120
Cell
A=B=C: 496.8 Å α=β=γ: 90.0 °
CCP4 map header:
mode
Image stored as Reals
Å/pix. X/Y/Z
4.14
4.14
4.14
M x/y/z
120
120
120
origin x/y/z
0.000
0.000
0.000
length x/y/z
496.800
496.800
496.800
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
-38
-19
-20
NX/NY/NZ
85
80
82
MAP C/R/S
1
2
3
start NC/NR/NS
-60
-60
-60
NC/NR/NS
120
120
120
D min/max/mean
-0.278
0.528
-0.001
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Supplemental data
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Sample components
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Entire : Filament of phosphofructokinase-1 liver isoform
Entire
Name: Filament of phosphofructokinase-1 liver isoform
Components
Complex: Filament of phosphofructokinase-1 liver isoform
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Supramolecule #1: Filament of phosphofructokinase-1 liver isoform
Supramolecule
Name: Filament of phosphofructokinase-1 liver isoform / type: complex / ID: 1 / Parent: 0 Details: PFKL tetramers assembled into filaments by addition of ATP and fructose-6-phosphate
Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 120 / Average exposure time: 1.2 sec. / Average electron dose: 30.0 e/Å2
Experimental equipment
Model: Tecnai Spirit / Image courtesy: FEI Company
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Image processing
Segment selection
Number selected: 9324 / Software - Name: Appion Helix Boxer
CTF correction
Software - Name: CTFFIND (ver. 3.0)
Startup model
Type of model: OTHER / Details: Featureless cylinder
Final angle assignment
Type: NOT APPLICABLE / Software - Name: SPIDER (ver. 21.00)
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