Examination of the Molecular Properties Underlying the Mechanism, Structure, and Specificity of LanB Enzymes Involved in Lanthipeptide Biosynthesis

检查参与羊毛硫肽生物合成的 LanB 酶的机制、结构和特异性的分子特性

• 批准号: 9525480
• 负责人: Ian Roger Bothwell
• 金额: $ 0.07万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-16 至 2019-03-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9525480
• 关键词: Active Sites; Address; Amides; Amino Acids; Amino Acyl Transfer RNA; Anabolism; Antibiotic Resistance; Antibiotics; Biochemical; Biomedical Engineering; Biomedical Research; Catalysis; Catalytic Domain; Chemicals; Communities; Complex; Cyclization; Cysteine; Data; Dehydration; Development; Diagnostic radiologic examination; Drug resistance; Enzyme Kinetics; Enzymes; Evolution; Food Industry; Foundations; Future; Generations; Glutamates; Glutamic Acid; Glutamic Acid-Specific tRNA; Goals; Health; Human; Hydro-Lyases; Isotope Labeling; Light; Mediating; Methods; Molecular; Mutagenesis; Names; Natural Products; Nature; Nisin; Peptides; Pharmacologic Substance; Process; Production; Property; Reaction; Reporting; Research; Resistance development; Ribosomes; Role; Serine; Side; Site; Specific qualifier value; Specificity; Stable Isotope Labeling; Structure; Sulfhydryl Compounds; Techniques; Therapeutic; Threonine; Transfer RNA; Work; analog; antimicrobial; antimicrobial drug; carboxylate; cofactor; combat; dehydroalanine; dehydrobutyrine; design; experimental study; hydroxyl group; insight; lanthionine; member; mutant; novel; pathogen; peptide structure; public health relevance

 DESCRIPTION (provided by applicant): One of the most pressing challenges to the biomedical research community lies in the development of novel antimicrobials as a means of combating the evolution of antibiotic-resistant pathogens. As such, research in the van der Donk is focused on the study of lanthipeptides, a class of ribosomally synthesized and posttranslationally modified peptide (RiPP) natural products, which have demonstrated significant potential as antibiotics. Indeed, the antibiotic lanthipeptide, nisin, has been used in the U.S. food industry for almost half a century with very few reports of antibiotic resistance. These natural products derive their name from the presence of lanthionine rings within their structure, which are formed through the conjugation of cysteine thiols to dehydroalanine and dehydrobutyrine residues within the maturing lanthipeptide. It has recently been found that LanBs, the enzymes responsible for the generation of dehydroalanine and dehydrobutyrine residues in some lanthipeptides, rely upon glutamyl-tRNAGlu in order to glutamylate and subsequently eliminate the hydroxyl-group from serine and threonine. Unfortunately, little is known about the enzymatic mechanisms by which this occurs or how it is regulated. Therefore, through the use of isotopically labeled cofactors, non-reactive substrate mimics, mutagenesis, and enzyme kinetics analysis, the research proposed herein is designed to examine the chemical and structural features that underlie LanB activity. The specific goals to be achieved include the determination of the mechanism and chemoselectivity of glutamate transfer by NisB, the structural elucidation of the glutamylation and elimination active sites, and the molecular mechanisms that determine tRNA cofactor recognition. Through a better understanding of these properties, our overall objective is to better understand lanthipeptide biosynthesis in order to inform the development of novel antibiotic and therapeutic compounds.

 描述（由适用提供）：生物医学研究界最紧迫的挑战之一在于开发新型抗微生物剂，作为打击抗生素耐药病原体进化的一种手段。因此，在范德多克（Van der Donk）中进行的研究集中在研究淋巴染的研究上，这是一类核糖体合成和翻译后修饰的肽（RIPP）天然产物的研究，这些肽（RIPP）天然产物具有抗生素的显着潜力。实际上，抗生素型灯lipeptide，Nisin已被用于 美国食品工业将近半个世纪的抗生素耐药性报道很少。这些天然产物的名称来自其结构内的灯笼环的存在，它们是通过半胱氨酸硫醇与脱氢丙氨酸和脱氢丁丁的结合而形成的。最近发现，负责产生脱氢丙氨酸和脱氢丁丁的酶在某些灯笼中保留，依靠谷氨酸 - trnaglu，以便从丝氨酸和他这里消除羟基。不幸的是，关于发生这种情况或如何调节的酶机制知之甚少。因此，通过使用同位素标记的辅因子，非反应性底物模拟，诱变和酶动力学分析，本文提出的研究旨在检查LANB活性的化学和结构特征。要实现的具体目标包括确定NISB通过NISB的机理和化学选择性的确定，谷氨酸的结构阐明和消除活性位点以及确定sTRNA辅助识别的分子机制。通过更好地理解这些特性，我们的总体目标是更好地了解植物肽生物合成，以便为新型抗生素和治疗化合物的发展提供信息。