National Natural Science Foundation of China (NSFC)
31971123
China
National Natural Science Foundation of China (NSFC)
81920108015
China
National Natural Science Foundation of China (NSFC)
31930059
China
Citation
Journal: Cell Discov / Year: 2021 Title: Mechanism of substrate transport and inhibition of the human LAT1-4F2hc amino acid transporter. Authors: Renhong Yan / Yaning Li / Jennifer Müller / Yuanyuan Zhang / Simon Singer / Lu Xia / Xinyue Zhong / Jürg Gertsch / Karl-Heinz Altmann / Qiang Zhou / Abstract: LAT1 (SLC7A5) is one of the representative light chain proteins of heteromeric amino acid transporters, forming a heterodimer with its heavy chain partner 4F2hc (SLC3A2). LAT1 is overexpressed in ...LAT1 (SLC7A5) is one of the representative light chain proteins of heteromeric amino acid transporters, forming a heterodimer with its heavy chain partner 4F2hc (SLC3A2). LAT1 is overexpressed in many types of tumors and mediates the transfer of drugs and hormones across the blood-brain barrier. Thus, LAT1 is considered as a drug target for cancer treatment and may be exploited for drug delivery into the brain. Here, we synthesized three potent inhibitors of human LAT1, which inhibit transport of leucine with IC values between 100 and 250 nM, and solved the cryo-EM structures of the corresponding LAT1-4F2hc complexes with these inhibitors bound at resolution of up to 2.7 or 2.8 Å. The protein assumes an outward-facing occluded conformation, with the inhibitors bound in the classical substrate binding pocket, but with their tails wedged between the substrate binding site and TM10 of LAT1. We also solved the complex structure of LAT1-4F2hc with 3,5-diiodo-L-tyrosine (Diiodo-Tyr) at 3.4 Å overall resolution, which revealed a different inhibition mechanism and might represent an intermediate conformation between the outward-facing occluded state mentioned above and the outward-open state. To our knowledge, this is the first time that the outward-facing conformation is revealed for the HAT family. Our results unveil more important insights into the working mechanisms of HATs and provide a structural basis for future drug design.
History
Deposition
Dec 31, 2020
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Header (metadata) release
Mar 10, 2021
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Map release
Mar 10, 2021
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Update
Jun 29, 2022
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Current status
Jun 29, 2022
Processing site: PDBj / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Download / File: emd_30836.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation
cryo EM map of the LAT1-4F2hc bound with JX-075, focused refined on transmembrane region
Voxel size
X=Y=Z: 1.087 Å
Density
Contour Level
By AUTHOR: 0.04 / Movie #1: 0.04
Minimum - Maximum
-0.09681396 - 0.18550214
Average (Standard dev.)
-0.00021625518 (±0.004469865)
Symmetry
Space group: 1
Details
EMDB XML:
Map geometry
Axis order
X
Y
Z
Origin
0
0
0
Dimensions
256
256
256
Spacing
256
256
256
Cell
A=B=C: 278.272 Å α=β=γ: 90.0 °
CCP4 map header:
mode
Image stored as Reals
Å/pix. X/Y/Z
1.087
1.087
1.087
M x/y/z
256
256
256
origin x/y/z
0.000
0.000
0.000
length x/y/z
278.272
278.272
278.272
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
0
0
0
NX/NY/NZ
288
288
288
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
0
NC/NR/NS
256
256
256
D min/max/mean
-0.097
0.186
-0.000
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Supplemental data
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Sample components
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Entire : cryo EM map of the LAT1-4F2hc bound with JX-075, focused refined ...
Entire
Name: cryo EM map of the LAT1-4F2hc bound with JX-075, focused refined on transmembrane region
Components
Complex: cryo EM map of the LAT1-4F2hc bound with JX-075, focused refined on transmembrane region
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Supramolecule #1: cryo EM map of the LAT1-4F2hc bound with JX-075, focused refined ...
Supramolecule
Name: cryo EM map of the LAT1-4F2hc bound with JX-075, focused refined on transmembrane region type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)
Organism: Homo sapiens (human)
Recombinant expression
Organism: Homo sapiens (human)
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Experimental details
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Structure determination
Method
cryo EM
Processing
single particle reconstruction
Aggregation state
particle
-
Sample preparation
Buffer
pH: 8
Vitrification
Cryogen name: ETHANE
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Electron microscopy
Microscope
FEI TITAN KRIOS
Image recording
Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron optics
Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
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