関連する実験動画

Updated: Jan 8, 2026

A Fluorescence-based Assay of Phospholipid Scramblase Activity

A Fluorescence-based Assay of Phospholipid Scramblase Activity

Published on: September 20, 2016

ドール鎖結合オリゴ糖のRft1を介したスクランブリングの提案メカニズム

George N Chiduza¹, Kentaro Sakata², Faria Noor²

¹Biochemistry and Structural Biology-Chemistry Department, Université Libre de Bruxelles - Campus Plaine, 1050 Brussels, Belgium.

bioRxiv : the preprint server for biology

|December 22, 2025

まとめ

酵母Rft1タンパク質、N結合型糖鎖中間体Man5GlcNAc2-PP-dolichol（M5-DLO）のスクランブラーゼは、交互アクセス機構を使用します。MurJとは異なり、Rft1は静電相互作用に柔軟性を示し、特殊な脂質輸送を示唆しています。

科学分野:

生化学; 分子生物学; 構造生物学

背景:

Rft1はER膜タンパク質であり、N結合型糖鎖形成に不可欠なMan5GlcNAc2-PP-dolichol（M5-DLO）のスクランブラーゼです。;Rft1はMOPトランスポータースーパーファミリーに属し、細菌のMurJリピドIIフリッパーゼと類似していますが、その輸送メカニズムは不明です。

研究の目的:

M5-DLOに対する酵母Rft1の輸送メカニズムを解明すること。;Rft1とMurJの機能的および構造的メカニズムを比較すること。

主な方法:

酵母Rft1-M5-DLO複合体のモデリングにAlphaFold3とChai1を利用しました。;Rft1空洞残基の比較変異分析を実施しました。;酵母レポーターストレインを用いてRft1機能を評価しました。

主要な成果:

構造モデルは、内向き開口状態、閉塞状態、外向き状態を伴うRft1の交互アクセス輸送メカニズムを示唆しています。;中央の空洞はM5-DLOのヘッド基を結合し、一方、ドルコールテールはポータルを介して疎水性溝に移動します。;Rft1は中央空洞における電荷反転変異に対して高い耐性を示し、MurJの厳密な静電要件から大きく逸脱しています。

結論:

Rft1はM5-DLOの特殊な交互アクセス輸送体として機能します。;Rft1機能に不可欠な主要な分子決定因子が特定されました。;交互アクセスメカニズムは、Rft1を脂質輸送に親水性溝を使用する既知のスクランブラーゼと区別します。

キーワード:

リピドII N結合型糖鎖交互アクセス機構先天性糖鎖形成不全症ドール鎖結合オリゴ糖小胞体ペプチドグリカンスクランブラーゼ

さらに関連する動画

Chemo-enzymatic Synthesis of N-glycans for Array Development and HIV Antibody Profiling

Chemo-enzymatic Synthesis of N-glycans for Array Development and HIV Antibody Profiling

Published on: February 5, 2018

Hierarchical and Programmable One-Pot Oligosaccharide Synthesis

Hierarchical and Programmable One-Pot Oligosaccharide Synthesis

Published on: September 6, 2019

関連する概念動画

Oligosaccharide Assembly

Oligosaccharide Assembly

Protein glycosylation starts in the ER lumen and continues in the Golgi apparatus. Glycosyltransferases catalyze the addition of sugar molecules or glycosylation of proteins. Usually, these enzymes add sugars to the hydroxyl groups of selected serine or threonine residues to form O-linked glycans or the amino groups of asparagine residues to form N-linked glycans. Different positions on the same polypeptide chain can contain differently linked glycans.
Multiple sugar molecules that may or may...

Membrane Asymmetry Regulating Transporters

Membrane Asymmetry Regulating Transporters

Enzymes like flippase, floppase, and scramblase transfer phospholipids from one layer to another in the membrane, thereby affecting membrane asymmetry.
Flippase
Eukaryotic flippases are type-IV P-type ATPases or P4-ATPases belonging to P-type ATPase family proteins that are membrane-bound pumps involved in the ATP-mediated transport of ions and molecules across the membrane. Flippases flip specific phospholipids from the outer to the inner leaflet of a membrane. All P4-ATPases have one...

Protein Modifications in the RER

Protein Modifications in the RER

Modification of secretory and transmembrane proteins entering the rough ER begins in the ER lumen. These modifications aid in protein folding and stabilize the acquired tertiary structure. Protein modifications in the rough ER co-occur at different stages of protein folding.
Broadly, these modifications can be categorized into four main categories — glycosylation, formation of disulfide bonds, assembly of protein subunits, and specific proteolytic cleavages like removal of signal...

Export of Misfolded Proteins out of the ER

Export of Misfolded Proteins out of the ER

After folding, the ER assesses the quality of secretory and membrane proteins. The correctly folded proteins are cleared by the calnexin cycle for transport to their final destination, while misfolded proteins are held back in the ER lumen. The ER chaperones attempt to unfold and refold the misfolded proteins but sometimes fail to achieve the correct native conformation. Such terminally misfolded proteins are then exported to the cytosol by ER-associated degradation or ERAD pathway for...

Protein Folding Quality Check in the RER

Protein Folding Quality Check in the RER

ER is the primary site for the maturation and folding of soluble and transmembrane secretory proteins. The calnexin cycle is a specific chaperone system that folds and assesses the confirmation of N-glycosylated proteins before they can exit the ER lumen. The primary players of this quality check pipeline are the lectins, ER-resident chaperones, and a glucosyl transferase enzyme. In case the calnexin system in the lumen fails to salvage a misfolded protein, it is transported to the cytoplasm...

Rab Cascades

Rab Cascades

Rab GTPases act in a regulated cascade during membrane fusion, helping the lipid bilayers mix. The Rab family of proteins are active when bound to GTP, and inactive when bound to GDP. Hence, they act as guanine nucleotide-dependent molecular switches. Rab-GTP recognizes and binds to long or short-range tethering proteins to capture the target vesicle. These tethers coordinate with SNAREs on the vesicle and the target membrane to assemble the trans SNARE complex that locks the mixing bilayers.