GST タグ融合タンパク質精製用クロマトグラフィー担体COSMOGEL® GST-Accept
GST(グルタチオン S-トランスフェラーゼ)タグは、タンパク質のアフィニティー精製において His タグと並び使用頻度の高いエピトープタグです。GST がそのリガンドであるグルタチオンと特異的に結合することから、グルタチオンを担体へ連結することで効率的に GST タグ融合タンパク質を精製することができます。
特長
- 優れた耐久性
- 優れたコストパフォーマンス
性能評価
GST タグ融合タンパク質の 10 回連続精製
GST-His を発現させた大腸菌ライセートを同一のゲルを用いて 10 回連続して精製しました。10 回目の精製においても問題なく精製できています。
- タンパク質マーカー(10倍濃縮)(#29458-24)を 1 × で使用
- 大腸菌ライセート
- 1 回目の溶出画分
- 2 回目の溶出画分
- 3 回目の溶出画分
- 4 回目の溶出画分
- 6 回目の溶出画分
- 8 回目の溶出画分
- 10 回目の溶出画分
- タンパク質マーカー(10倍濃縮)(#29458-24)を 1 × で使用
サンプル:E. coli(BL21(DE3)pLysS) lysate transformed with GST-His(pET-41b(+))
結合緩衝液:Phosphate Buffered Saline(10x)(pH 7.4)(#27575-31)を 1 × で使用
溶出緩衝液:10 mM Glutathione, 50 mM Tris-HCl, pH 8.0
使用例
GST タグ融合タンパク質の Thrombin によるカラム内消化
Thrombin(#33842-44)無添加の場合は発現タンパク質がそのまま回収されていますが、Thrombin 添加の場合は PBS 画分(Lane7)に S タグ - His タグが溶出しています。このことから、カラム内消化も問題なく行えていることがわかります。
- タンパク質マーカー(10倍濃縮)(#29458-24)を 1 × で使用
- 大腸菌ライセート
- フロースルー画分
- Thrombin (-) PBS fraction
- Thrombin (-) glutathione fraction
- フロースルー画分
- Thrombin (+) PBS fraction
- Thrombin (+) glutathione fraction
- 大腸菌ライセート
- タンパク質マーカー(10倍濃縮)(#29458-24)を 1 × で使用
*1: GST タグ - His タグ - S タグ - His タグ
*2: S タグ - His タグ
*3: GST タグ - His タグ
サンプル:E. coli(BL21(DE3)pLysS) lysate transformed with GST-His(pET-41b(+))
結合緩衝液:Phosphate Buffered Saline(10x)(pH 7.4)(#27575-31)を 1 × で使用
溶出緩衝液:10 mM Glutathione, 50 mM Tris-HCl, pH 8.0
薬品耐性
回収量に影響を及ぼさない添加物・緩衝液
還元剤 | 20 mmol/L DTT | 20 mmol/L 2-Mercaptoethanol |
---|---|---|
20 mmol/L TCEP | ||
界面活性剤 | 1% Triton X-100 | 1% Tween 20 |
1% Nonidet P-40 | 1% CHAPS | |
1% Brij 35 | 1% 1-O-n-Octyl-β-D-glucopyranoside |
|
1% n-Dodecyl-β-D-maltopyranoside |
||
添加剤 | 50 mmol/L EDTA | 50 mmol/L EGTA |
2 mol/L NaCl | 1 mol/L MgCl2 | |
500 mmol/L Imidazole | 20% Ethanol | |
Protease Inhibitor Cocktail※1 (5×) |
Phosphatase Inhibitor Cocktail※2 (5×) |
|
緩衝液 | 100 mmol/L HEPES(pH 7.4, 8.0) | 100 mmol/L Tris-HCl(pH 7.4, 8.0) |
上記添加物を含む結合・溶出緩衝液を用いて確認したものであり、上限値を示すものではありません。また記載されていない添加物の使用を制限するものでもありません。融合している標的タンパク質によっては上記添加物の影響を受ける場合がありますので、あらかじめ小スケールで検討を行ってください。
※1 弊社 Protease Inhibitor Cocktail(100×)(一般用:#04080、EDTA free:#03969)
※2 弊社 Phosphatase Inhibitor Cocktail(100×)(#07574-61、EDTA free:#07575-51)
Triton はユニオン・カーバイド・コーポレーション、Tween はクローダ インターナショナル ピーエルシー、Nonidet はシェル ブランズ インターナショナル アクチェンゲゼルシャフト、Brij はクローダ アメリカス リミテッド ライアビリティ カンパニーの登録商標です。
製品仕様
リガンド | グルタチオン |
---|---|
粒子サイズ | 50 ~ 150 µm |
マトリックス | 4% アガロース |
回収容量 | >5 mg/mL(回収量 / ゲル量) |
推奨線流速 | 26 cm/h |
限界圧 | 2.6 psi(0.18 bar) |
供給状態 | 50vol% 懸濁液(20vol% エタノール水溶液) |
使用文献
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価格表
※掲載内容は予告なく変更になる場合があります。