大腸桿菌tRNA的中文介紹  
大腸桿菌（*Escherichia coli*）的轉(zhuǎn)運(yùn)RNA（tRNA）是蛋白質(zhì)合成的核心分子，負(fù)責(zé)將氨基酸精準(zhǔn)傳遞至核糖體，并根據(jù)mRNA的密碼子序列組裝多肽鏈。以下從結(jié)構(gòu)、加工、基因組織及功能調(diào)控等方面詳述其特性：  

1. **分子結(jié)構(gòu)與功能**  
- **二級結(jié)構(gòu)**：tRNA呈典型的三葉草結(jié)構(gòu)，包含四個(gè)功能區(qū)域：接受莖（3'端攜帶氨基酸）、D環(huán)、反密碼子環(huán)（含反密碼子）和TΨC環(huán)。反密碼子環(huán)中的三核苷酸序列（反密碼子）與mRNA上的密碼子互補(bǔ)配對，確保氨基酸的精準(zhǔn)定位。  
- **接受莖與氨基?；?*：接受莖的3'末端通過高能酯鍵連接特定氨基酸，這一過程由氨基酰-tRNA合成酶（AARS）催化。AARS分為I類和II類，分別將氨基酸連接到tRNA的2'-OH或3'-OH位，但溶液中兩者可自發(fā)異構(gòu)化。  

2. **tRNA的加工過程**  
- **前體轉(zhuǎn)錄與切割**：大腸桿菌的tRNA基因大多以多順反子形式存在，與rRNA或蛋白質(zhì)基因共轉(zhuǎn)錄。前體tRNA（pre-tRNA）需經(jīng)歷多步加工：  
  1. **5'端切割**：由核酶RNase P（含RNA亞基M1和蛋白質(zhì)C5）切除5'前導(dǎo)序列。  
  2. **3'端修剪**：通過內(nèi)切酶（如RNase E）和外切酶（如RNase T）逐步切除3'尾隨序列，最終暴露或添加CCA序列。約30%的tRNA需通過tRNA核苷酸轉(zhuǎn)移酶在3'端后加CCA，該酶無需模板即可依次添加C、C、A。  
- **堿基修飾**：包括甲基化、硫代化等，增強(qiáng)tRNA穩(wěn)定性和功能特異性。  

3. **基因組織與表達(dá)調(diào)控**  
- **基因分布**：大腸桿菌基因組含86個(gè)tRNA基因，編碼40種成熟tRNA異型（對應(yīng)20種氨基酸）。其中64個(gè)基因位于多順反子轉(zhuǎn)錄單元，常與rRNA或蛋白基因共存；其余為單順反子。  
- **動態(tài)調(diào)控**：在氨基酸饑餓條件下，tRNA的穩(wěn)定性顯著下降，20分鐘內(nèi)大部分被降解，以快速響應(yīng)代謝壓力。此外，多腺苷酸化酶（PAP I）通過在前體tRNA的3'端添加短腺苷酸尾，與外切酶競爭，調(diào)控tRNA的成熟與穩(wěn)定性。  

4. **研究案例與技術(shù)進(jìn)展**  
- **亮氨酸t(yī)RNA（tRNA^Leu^）** ：通過優(yōu)化提取與純化方法，研究者獲得高純度tRNA^Leu^，其3'和5'端存在額外核苷酸，且能特異性結(jié)合亮氨酸。  
- **基因工程應(yīng)用**：利用質(zhì)粒pTrc99B-Leu1/Leu2在大腸桿菌中高效表達(dá)tRNA^Leu^，為研究tRNA與合成酶的相互作用提供材料。  

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Introduction to *Escherichia coli* tRNA (English Version)  
The transfer RNA (tRNA) of *Escherichia coli* serves as a molecular adaptor that links mRNA codons to amino acids during protein synthesis. Below is a comprehensive overview of its structure, biogenesis, and functional regulation:  

1. **Molecular Architecture and Functional Domains**  
- **Secondary Structure**: tRNA adopts a cloverleaf-like fold with four domains: the acceptor stem (3' end for amino acid attachment), D-loop, anticodon loop (containing the anticodon triplet), and TΨC loop. The anticodon pairs with mRNA codons via complementary base pairing.  
- **Aminoacylation**: Aminoacyl-tRNA synthetases (AARSs) catalyze the attachment of specific amino acids to the 3'-OH or 2'-OH of the terminal ribose in the acceptor stem. Class I and II AARSs differ in their binding modes, but spontaneous transacylation between 2' and 3' isomers occurs in solution.  

2. **tRNA Maturation and Processing**  
- **Precursor Cleavage**: Most tRNA genes are transcribed as polycistronic units alongside rRNA or protein-coding genes. Key processing steps include:  
  1. **5' End Trimming**: Catalyzed by RNase P, a ribozyme composed of an RNA subunit (M1) and a protein subunit (C5).  
  2. **3' End Maturation**: Achieved via endonucleases (e.g., RNase E) and exonucleases (e.g., RNase T). For some tRNAs, the 3' CCA sequence is added post-transcriptionally by tRNA nucleotidyltransferase, which sequentially incorporates CTP, CTP, and ATP without a template.  
- **Nucleotide Modifications**: Chemical modifications (e.g., methylation, thiolation) enhance tRNA stability and decoding accuracy.  

3. **Genomic Organization and Regulatory Mechanisms**  
- **Gene Clusters**: The *E. coli* genome harbors 86 tRNA genes encoding 40 mature isoacceptors. Approximately 75% reside in polycistronic operons, while others are monocistronic.  
- **Dynamic Regulation**: Under amino acid starvation, tRNAs undergo rapid degradation within 20 minutes, reflecting their role in metabolic adaptation. Polyadenylation by PAP I competes with 3'→5' exonucleases, modulating tRNA stability and functional availability.  

4. **Research Advances and Applications**  
- **Leucine tRNA (tRNA^Leu^)**: High-purity tRNA^Leu^ was isolated with unique terminal nucleotides, demonstrating specific leucine-binding capacity.  
- **Genetic Engineering**: Plasmid-based overexpression systems (e.g., pTrc99B-Leu1/Leu2) enable large-scale production of tRNA^Leu^, facilitating studies on tRNA-synthetase interactions.  

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