National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)
R01AI147890
米国
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)
5U54AI150472-09
米国
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)
U54 AI170855-01
米国
Medical Research Council (MRC, United Kingdom)
MC_UP_1201/16
英国
Max Planck Society
ドイツ
Other private
RRID:SCR_021727
Other private
RRID:SCR_021740
National Science Foundation (NSF, United States)
DMR-1719875
米国
引用
ジャーナル: Proc Natl Acad Sci U S A / 年: 2023 タイトル: Structural insights into HIV-1 polyanion-dependent capsid lattice formation revealed by single particle cryo-EM. 著者: Carolyn M Highland / Aaron Tan / Clifton L Ricaña / John A G Briggs / Robert A Dick / 要旨: The HIV-1 capsid houses the viral genome and interacts extensively with host cell proteins throughout the viral life cycle. It is composed of capsid protein (CA), which assembles into a conical ...The HIV-1 capsid houses the viral genome and interacts extensively with host cell proteins throughout the viral life cycle. It is composed of capsid protein (CA), which assembles into a conical fullerene lattice composed of roughly 200 CA hexamers and 12 CA pentamers. Previous structural analyses of individual CA hexamers and pentamers have provided valuable insight into capsid structure and function, but detailed structural information about these assemblies in the broader context of the capsid lattice is lacking. In this study, we combined cryoelectron tomography and single particle analysis (SPA) cryoelectron microscopy to determine structures of continuous regions of the capsid lattice containing both hexamers and pentamers. We also developed a method of liposome scaffold-based in vitro lattice assembly ("lattice templating") that enabled us to directly study the lattice under a wider range of conditions than has previously been possible. Using this approach, we identified a critical role for inositol hexakisphosphate in pentamer formation and determined the structure of the CA lattice bound to the capsid-targeting antiretroviral drug GS-6207 (lenacapavir). Our work reveals key structural details of the mature HIV-1 CA lattice and establishes the combination of lattice templating and SPA as a robust strategy for studying retroviral capsid structure and capsid interactions with host proteins and antiviral compounds.
全体 : HIV-1 CA protein capsid-like particles assembled in the presence ...
全体
名称: HIV-1 CA protein capsid-like particles assembled in the presence of inositol hexakisphosphate
要素
複合体: HIV-1 CA protein capsid-like particles assembled in the presence of inositol hexakisphosphate
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超分子 #1: HIV-1 CA protein capsid-like particles assembled in the presence ...
超分子
名称: HIV-1 CA protein capsid-like particles assembled in the presence of inositol hexakisphosphate タイプ: complex / ID: 1 / キメラ: Yes / 親要素: 0
由来(天然)
生物種: Escherichia coli (大腸菌)
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実験情報
-
構造解析
手法
クライオ電子顕微鏡法
解析
サブトモグラム平均法
試料の集合状態
particle
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試料調製
緩衝液
pH: 6.2 構成要素:
濃度
式
名称
50.0 mM
MES
MES
2.5 mM
C6H18O24P6
Inositol hexakisphosphate
1.0 mM
C9H15O6P
Tris(2-carboxyethyl)phosphine
グリッド
モデル: C-flat-2/2 / 材質: COPPER / メッシュ: 300 / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 時間: 30 sec. / 前処理 - 雰囲気: AIR 詳細: The grid was glow discharged for 30 seconds at a current of 20 mA.
凍結
凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 277.15 K / 装置: FEI VITROBOT MARK IV
想定した対称性 - 点群: C5 (5回回転対称) / アルゴリズム: BACK PROJECTION / 解像度のタイプ: BY AUTHOR / 解像度: 6.2 Å / 解像度の算出法: FSC 0.143 CUT-OFF / ソフトウェア - 名称: subTOM / 使用したサブトモグラム数: 5244
抽出
トモグラム数: 66 / 使用した粒子像数: 5264 / ソフトウェア - 名称: MATLAB 詳細: Pentamer positions in HIV-1 CA protein capsid-like particles were identified by identifying patterns of 5 aligned hexamer positions corresponding to pentamer geometry using MATLAB.