設計されたフェニララニル-tRNA合成酵素の変種は,アリルケトンの機能がタンパク質に効率的に組み込まれることをin vivoで可能にします
Deepshikha Datta1, Pin Wang, Isaac S Carrico
1Division of Chemistry and Chemical Engineering, Howard Hughes Medical Institute, California Institute of Technology, Pasadena, California 91125, USA.
Journal of the American Chemical Society
|May 16, 2002
まとめ
研究者は非天然のアミノ酸を組み込むことでタンパク質合成を拡大しました. 彼らは突然変異した酵素を使用して,dihydrofolate reductaseにp-acetylphenylalanineを添加し,化学結合を可能にしました.
科学分野:
- バイオケミストリーと分子生物学
- 合成生物学 合成生物学とは
- プロテイン工学は,タンパク質の
背景:
- タンパク質合成のために利用可能なアミノ酸のレパートリーを拡大することは,タンパク質の設計と機能を進めるために極めて重要です.
- 非自然なアミノ酸をインビヴォに組み込むことは,新しいタンパク質の機能を生み出すための強力な戦略を提供します.
- 化学選択性結合により,穏やかな条件下でタンパク質の正確な改変が可能になります.
研究 の 目的:
- 非天然のアミノ酸であるp-アセチルフェニララニンが,再結合タンパク質にインビボ組み込まれることを実証する.
- 非自然なアミノ酸を効率的に組み込むために宿主生物のアミノアシル-tRNA合成酵素を設計する.
- 添加された非自然なアミノ酸を,その後の化学選択タンパク質結合に利用する.
主な方法:
- エシェリキア・コロイ菌からの変異したフェニララニル-tRNA合成酵素の計算設計.
- 活体内でのp-アセチルフェニララニンの組み込みは,再結合ダイヒドロフォラート還元酵素 (DHFR) に.
- ケトン機能化されたDHFRをヒドラジド反応剤で化学選択的に結合する.
主要な成果:
- p-アセチルフェニララニンをE. coliの再結合DHFRに成功裏に組み込んだ.
- エンジニアリングされたフェニララニル-tRNA合成酵素は,非自然なアミノ酸の特定の充電を促進しました.
- DHFRのケトン機能は,穏やかな条件下で効率的な化学選択結合を可能にしました.
結論:
- この研究では,ケトン機能化されたアミノ酸をタンパク質にインビヴォで組み込む方法が確立されています.
- 開発されたアプローチは,タンパク質合成と設計のためのツールキットを拡張し,新しい機能化戦略を可能にします.
- エンジニアリングされたタンパク質の化学選択的結合は,生物結合とタンパク質の改変のための多用途のプラットフォームを提供します.
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関連する概念動画
RNA Structure
Overview
The basic structure of RNA consists of a five-carbon sugar and one of four nitrogenous bases. Although most RNA is single-stranded, it can form complex secondary and tertiary structures. Such structures play essential roles in the regulation of transcription and translation.
Different Types of RNA Have the Same Basic Structure
There are three main types of ribonucleic acid (RNA): messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA). All three RNA types consist of a...
The basic structure of RNA consists of a five-carbon sugar and one of four nitrogenous bases. Although most RNA is single-stranded, it can form complex secondary and tertiary structures. Such structures play essential roles in the regulation of transcription and translation.
Different Types of RNA Have the Same Basic Structure
There are three main types of ribonucleic acid (RNA): messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA). All three RNA types consist of a...
Transfer RNA Synthesis
One of the unique features of tRNA is the presence of modified bases. In some tRNAs, modified bases account for nearly 20% of the total bases in the molecule. Altogether, these unusual bases protect the tRNA from enzymatic degradation by RNases.
Each of these chemical modifications is carried by a specific enzyme, post-transcription. All of these enzymes have unique base and site-specificity. Methylation, the most common chemical modification, is carried by at least nine different enzymes, with...
Each of these chemical modifications is carried by a specific enzyme, post-transcription. All of these enzymes have unique base and site-specificity. Methylation, the most common chemical modification, is carried by at least nine different enzymes, with...
tRNA Activation
Aminoacyl-tRNA synthetases are present in both eukaryotes and bacteria. Though eukaryotes have 20 different aminoacyl-tRNA synthetases to couple to 20 amino acids, many bacteria do not have genes for all of these aminoacyl-tRNA synthetases. Despite this, they still use all 20 amino acids to synthesize their proteins. For instance, some bacteria do not have the gene encoding the enzyme that couples glutamine with its partner tRNA. In these organisms, one enzyme adds glutamic acid to all of the...
RNA Structure
The basic structure of RNA consists of a string of ribonucleotides attached by phosphodiester bonds. Although most RNA is single-stranded, it can form complex secondary and tertiary structures. Such structures play essential roles in the regulation of transcription and translation.
Different Types of RNA Have the Same Basic Structure
There are three main types of ribonucleic acid (RNA) involved in protein synthesis: messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA). All three...
Different Types of RNA Have the Same Basic Structure
There are three main types of ribonucleic acid (RNA) involved in protein synthesis: messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA). All three...
Transfer RNA Synthesis
One of the unique features of tRNA is the presence of modified bases. In some tRNAs, modified bases account for nearly 20% of the total bases in the molecule. Altogether, these unusual bases protect the tRNA from enzymatic degradation by RNases.
Each of these chemical modifications is carried by a specific enzyme, post-transcription. All of these enzymes have unique base and site-specificity. Methylation, the most common chemical modification, is carried by at least nine different enzymes, with...
Each of these chemical modifications is carried by a specific enzyme, post-transcription. All of these enzymes have unique base and site-specificity. Methylation, the most common chemical modification, is carried by at least nine different enzymes, with...
tRNA Activation
Aminoacyl-tRNA synthetases are present in both eukaryotes and bacteria. Though eukaryotes have 20 different aminoacyl-tRNA synthetases to couple to 20 amino acids, many bacteria do not have genes for all of these aminoacyl-tRNA synthetases. Despite this, they still use all 20 amino acids to synthesize their proteins. For instance, some bacteria do not have the gene encoding the enzyme that couples glutamine with its partner tRNA. In these organisms, one enzyme adds glutamic acid to all of the...
