FUNCTIONS OF tRNA MODIFICATIONS

tRNA 修饰的功能

• 批准号: 7582509
• 负责人: Ya-Ming Hou
• 金额: $ 30.9万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-15 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7582509
• 关键词: Active Sites; Address; Anabolism; Anticodon; Archaea; Bacteria; Binding; Biochemical; Catalysis; Cleaved cell; Communication; Complement; Complex; Coupled; Dissection; Drug Delivery Systems; Drug Design; Ensure; Enzymatic Biochemistry; Enzymes; Escherichia coli; Eukaryota; Eukaryotic Cell; Foundations; Frequencies; Genes; Genetic; Genetic Transcription; Goals; Growth; Health; Healthcare; Human; Kinetics; Life; Methionine; Modification; Molecular; Molecular Conformation; Monitor; Movement; Nucleic Acids; Pharmaceutical Preparations; Positioning Attribute; Preparation; Process; Protein Biosynthesis; Proteins; Public Health; Purpose; Rate; Reaction; Reading Frames; Resources; Ribosomes; Role; S-Adenosylmethionine; Salmonella typhimurium; Side; Site; Solid; Specificity; Streptococcus pneumoniae; Structure; Testing; Transfer RNA; Transferase; Translating; Vertebral column; analog; base; cell growth; cofactor; fitness; improved; insight; interest; methyl group; pathogen; research study; success; tool

DESCRIPTION (provided by applicant): Modifications of transfer RNA (tRNA) molecules occur after transcription and constitute an essential step to promote cellular fitness and viability. Of interest is the m1G37 modification, which takes place at almost all tRNA molecules that have the G37 base immediately adjacent to the 3' side of the anticodon sequence. The m1G37 modification is important for maintaining the tRNA reading frame specificity on the ribosome during protein synthesis. Elimination of this modification increases ribosomal errors in protein synthesis and elevates frequencies of frame shifts. The enzyme that catalyzes the m1G37 modification is tRNA(m1G37) methyl transferase, which transfers the methyl group of S-adenosyl methionine (AdoMet) to the N1 position of G37 in tRNA. The bacterial enzyme, known as TrmD (encoded by the trmD gene), is essential for growth in E. coli, Salmonella typhimurium, and Streptococcus pneumoniae. An unexpected recent finding is that while TrmD is highly conserved among bacterial species, it shares little sequence or structural homology with its eukaryotic and archaeal counterpart (known as Trm5, encoded by the trm5 gene). This establishes TrmD and Trm5 as a pair of analogous enzymes that use distinct structural folds to catalyze the same reaction to synthesize the growth-dependent m1G37 in tRNA. The separation of TrmD and Trm5 along the split of bacteria from eukarya- archaea thus raises the medically relevant and attractive prospect of selective targeting of the bacterial TrmD. This proposal is aimed at providing a strong biochemical and molecular foundation that is necessary for the success of such drug targeting. Two aims of the project are to determine the molecular and structural basis of TrmD and Trm5 for their recognition of AdoMet and tRNA and the dynamic recognition process that leads to catalysis. The third aim will determine the role of the m1G37 modification on the ribosome during the decoding process. Together, these aims combine the strengths and interests of two presently separate fields (enzymology of tRNA modification, and ribosome structure and function) to address key issues that will have long-term impact on human health and bio-defense against bacterial pathogens. PUBLIC HEALTH RELEVANCE: The cellular protein synthesis machinery provides the basis for translating genetic information stored in nucleic acids to functional proteins. This machinery depends on extensive tRNA-ribosome interactions to ensure the fidelity of protein synthesis. The m1G37 modification of tRNA is a key determinant of this fidelity and is essential for growth for several bacterial pathogens. However, the molecular and biochemical basis for how the m1G37 modification is synthesized and how it functions on the ribosome is poorly understood. Preliminary studies show that the enzyme that catalyzes the m1G37 synthesis in bacteria was of a different structural origin from its counterpart in eukaryotes and in archaea. This proposal is aimed at elucidating the molecular basis for the enzymatic synthesis of m1G37 and the differences between the bacterial and eukaryotic/archaeal enzymes. Information to be gained from this proposal will provide important insights to build a solid foundation for selective inhibition of the bacterial enzyme so as to improve human health care.

描述（由申请人提供）：转移RNA（tRNA）分子的修饰发生在转录后，并构成促进细胞适应性和活力的重要步骤。令人感兴趣的是 m1G37 修饰，几乎所有 G37 碱基紧邻反密码子序列 3' 侧的 tRNA 分子都会发生这种修饰。 m1G37 修饰对于在蛋白质合成过程中维持核糖体上 tRNA 阅读框的特异性非常重要。消除这种修饰会增加蛋白质合成中的核糖体错误并提高移码频率。催化 m1G37 修饰的酶是 tRNA(m1G37) 甲基转移酶，它将 S-腺苷甲硫氨酸 (AdoMet) 的甲基转移到 tRNA 中 G37 的 N1 位。这种细菌酶称为 TrmD（由 trmD 基因编码），对于大肠杆菌、鼠伤寒沙门氏菌和肺炎链球菌的生长至关重要。最近的一个意外发现是，虽然 TrmD 在细菌物种中高度保守，但它与其真核和古细菌对应物（称为 Trm5，由 trm5 基因编码）几乎没有序列或结构同源性。这将 TrmD 和 Trm5 确立为一对类似的酶，它们使用不同的结构折叠来催化相同的反应，以合成 tRNA 中的生长依赖性 m1G37。因此，TrmD 和 Trm5 沿着细菌从真核古细菌的分裂中分离，提出了选择性靶向细菌 TrmD 的医学相关和有吸引力的前景。该提案旨在提供此类药物靶向成功所必需的强大的生化和分子基础。该项目的两个目标是确定 TrmD 和 Trm5 识别 AdoMet 和 tRNA 的分子和结构基础以及导致催化的动态识别过程。第三个目标是确定 m1G37 修饰在解码过程中对核糖体的作用。这些目标结合了两个目前独立领域（tRNA 修饰的酶学以及核糖体结构和功能）的优势和利益，以解决对人类健康和细菌病原体生物防御产生长期影响的关键问题。公共健康相关性：细胞蛋白质合成机制为将核酸中存储的遗传信息转化为功能蛋白质提供了基础。该机制依赖于广泛的 tRNA-核糖体相互作用来确保蛋白质合成的保真度。 tRNA 的 m1G37 修饰是这种保真度的关键决定因素，并且对于多种细菌病原体的生长至关重要。然而，人们对 m1G37 修饰如何合成以及它如何在核糖体上发挥作用的分子和生化基础知之甚少。初步研究表明，细菌中催化 m1G37 合成的酶与真核生物和古细菌中的对应酶具有不同的结构起源。该提案旨在阐明 m1G37 酶促合成的分子基础以及细菌和真核/古细菌酶之间的差异。从该提案中获得的信息将为选择性抑制细菌酶奠定坚实的基础提供重要的见解，从而改善人类健康保健。