ジャーナル: Cell Rep / 年: 2017 タイトル: Conformational Freedom of the LRP6 Ectodomain Is Regulated by N-glycosylation and the Binding of the Wnt Antagonist Dkk1. 著者: Kyoko Matoba / Emiko Mihara / Keiko Tamura-Kawakami / Naoyuki Miyazaki / Shintaro Maeda / Hidenori Hirai / Samuel Thompson / Kenji Iwasaki / Junichi Takagi / 要旨: LDL-receptor-related protein 6 (LRP6) is a single-pass membrane glycoprotein with a large modular ectodomain and forms a higher order signaling platform upon binding Wnt ligands on the cell surface. ...LDL-receptor-related protein 6 (LRP6) is a single-pass membrane glycoprotein with a large modular ectodomain and forms a higher order signaling platform upon binding Wnt ligands on the cell surface. Although multiple crystal structures are available for fragments of the LRP6 ectodomain, we lack a consensus view on the overall molecular architecture of the full-length LRP6 and its dynamic aspects. Here, we used negative-stain electron microscopy to probe conformational states of the entire ectodomain of LRP6 in solution and found that the four-module ectodomain undergoes a large bending motion hinged at the junction between the second and the third modules. Importantly, the extent of inter-domain motion is modulated by evolutionarily conserved N-glycan chains proximal to the joint. We also found that the LRP6 ectodomain becomes highly compact upon complexation with the Wnt antagonist Dkk1, suggesting a potential role for the ectodomain conformational change in the regulation of receptor oligomerization and signaling.
ダウンロード / ファイル: emd_9501.map.gz / 形式: CCP4 / 大きさ: 30.5 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
ボクセルのサイズ
X=Y=Z: 2.2 Å
密度
表面レベル
登録者による: 0.198 / ムービー #1: 0.3
最小 - 最大
-0.079741 - 1.0378006
平均 (標準偏差)
0.00088656624 (±0.036129534)
対称性
空間群: 1
詳細
EMDB XML:
マップ形状
Axis order
X
Y
Z
Origin
0
0
0
サイズ
200
200
200
Spacing
200
200
200
セル
A=B=C: 440.0 Å α=β=γ: 90.0 °
CCP4マップ ヘッダ情報:
mode
Image stored as Reals
Å/pix. X/Y/Z
2.2
2.2
2.2
M x/y/z
200
200
200
origin x/y/z
0.000
0.000
0.000
length x/y/z
440.000
440.000
440.000
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
-190
-190
-190
NX/NY/NZ
380
380
380
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
0
NC/NR/NS
200
200
200
D min/max/mean
-0.080
1.038
0.001
-
添付データ
-
試料の構成要素
+
全体 : Human LRP6 ectodomain complexed with full length of Dkk1
全体
名称: Human LRP6 ectodomain complexed with full length of Dkk1
要素
複合体: Human LRP6 ectodomain complexed with full length of Dkk1
複合体: LRP6 ectodomain
タンパク質・ペプチド: Low-density lipoprotein receptor-related protein 6
タンパク質・ペプチド: Low-density lipoprotein receptor-related protein 6
複合体: Human Dickkopf-related protein1
タンパク質・ペプチド: Dickkopf-related protein 1
リガンド: N-ACETYL-D-GLUCOSAMINE
リガンド: PHOSPHATE ION
リガンド: ALPHA-L-FUCOSE
リガンド: GLYCEROL
リガンド: water
+
超分子 #1: Human LRP6 ectodomain complexed with full length of Dkk1
超分子
名称: Human LRP6 ectodomain complexed with full length of Dkk1 タイプ: complex / ID: 1 / 親要素: 0 / 含まれる分子: #1-#3 詳細: Human Dkk1 fused with human growth hormone at the N-terminal (hGH-Dkk1) was co-expressed with LRP6ec in the HEK293S GnT1- cells.
照射モード: SPOT SCAN / 撮影モード: BRIGHT FIELD / Cs: 2.8 mm / 倍率(公称値): 80000
試料ステージ
試料ホルダーモデル: SIDE ENTRY, EUCENTRIC
+
画像解析
詳細
The selected images were band-pass filtered.
粒子像選択
詳細: 10974 tilt pairs for RCT
CTF補正
ソフトウェア - 名称: spider 詳細: This was not done for RCT but for following single-particle reconstruction
初期モデル
モデルのタイプ: RANDOM CONICAL TILT / Random conical tilt - Number images: 10974
最終 再構成
想定した対称性 - 点群: C1 (非対称) / アルゴリズム: BACK PROJECTION / 解像度のタイプ: BY AUTHOR / 解像度: 21.0 Å / 解像度の算出法: FSC 0.5 CUT-OFF / ソフトウェア - 名称: spider / 使用した粒子像数: 5390
初期 角度割当
タイプ: PROJECTION MATCHING Projection matching processing - Number reference projections: 83 ソフトウェア - 名称: spider 詳細: The projections generated from RCT were used for initial angle assignment