Mechanism and Inhibition for LuxS: A Biodefense Target

LuxS 的机制和抑制：生物防御目标

• 批准号: 6719230
• 负责人: ZHAOHUI SUNNY ZHOU
• 金额: $ 30.07万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6719230
• 关键词: X ray crystallography; aldehyde lyase; analog; antibacterial agents; bacteria infection mechanism; bacterial toxins; biofilm; bioterrorism /chemical warfare; chemical synthesis; drug resistance; enzyme activity; enzyme inhibitors; enzyme substrate; high performance liquid chromatography; isomerase; ketones; lyase; mass spectrometry; quorum sensing; synthetic enzyme; virulence

DESCRIPTION (provided by applicant): Bacterial infections, particularly those emerging with antibiotic resistance, pose an alarming threat to public health. Our long-term objective is to identify, characterize and validate new antibacterial targets. Traditional antibiotics act by killing or inhibiting bacteria, hence inducing antibiotic resistance. It is therefore imperative to explore alternative or complementary approaches. In the past few years, the ubiquitous bacterial enzyme LuxS has been found to play diverse and pivotal roles in bacterial quorum sensing, virulence regulation, toxin secretion and biofilm formation. In addition, central metabolic roles for LuxS are also proposed. This enzyme is found in Category A pathogens, including B. anthracis and Yersinia pestis; and in Category B pathogens, including Vibrio cholerae, Salmonella and diarrheagenic E. coil. Absent in humans, LuxS is an attractive target for anti-infective agent development. The enzymatic mechanism of LuxS remains elusive. Based on our preliminary studies, we propose that LuxS possesses functions of both an aldose-ketose isomerase and a lyase. The dual function of LuxS is mechanistically intriguing. Our proposed mechanism involves an initial aldose-ketose isomerization to generate a ketone at the C3 position on the carbohydrate moiety, and a final beta-elimination to cleave the C-S bond in S-ribosylhomocysteine. Our Specific Aim 1 is to chemically synthesize the proposed intermediates and their analogs, and test them as LuxS substrates or inhibitors. We will also attempt to trap or directly observe the proposed intermediates. Our Specific Aims 2 and 3 are to investigate the catalytic roles of Glu57 and Cys84 in B. subtilis LuxS by mutagenesis and chemical rescue, and the biological relevance of Cys84 oxidation. Lastly, we plan to design, synthesize and test mechanism-based inhibitors for LuxS, particularly those interacting with the active site zinc ion. We will also investigate how halogenated furanones, a group of natural antibacterial agents, inactivates LuxS, particularly for the V. cholerae enzyme. Additionally, we will test the effects of LuxS inhibitors on quorum sensing, biofilm formation and related bacterial physiology.

描述（由申请人提供）：细菌感染，特别是那些出现抗生素耐药性的细菌感染，对公共卫生构成了令人担忧的威胁。我们的长期目标是识别、表征和验证新的抗菌靶标。传统抗生素通过杀死或抑制细菌起作用，从而诱导抗生素耐药性。因此，必须探索替代或补充办法。在过去的几年中，普遍存在的细菌酶LuxS已被发现在细菌群体感应、毒力调节、毒素分泌和生物膜形成中发挥多种和关键的作用。此外，还提出了LuxS的中心代谢作用。这种酶存在于A类病原体中，包括B类。炭疽和鼠疫耶尔森氏菌;以及B类病原体，包括霍乱弧菌、沙门氏菌和大肠杆菌。线圈由于在人类中不存在，LuxS是抗感染药物开发的一个有吸引力的靶标。 LuxS的酶促机制仍然难以捉摸。根据我们的初步研究，我们认为LuxS同时具有醛糖-酮糖异构酶和裂解酶的功能。LuxS的双重功能在机制上是有趣的。我们提出的机制涉及初始的醛糖-酮糖异构化，以在碳水化合物部分的C3位置产生酮，以及最终的β-消除，以切割S-核糖基高半胱氨酸中的C-S键。我们的具体目标1是化学合成所提出的中间体及其类似物，并测试它们作为LuxS底物或抑制剂。我们还将尝试捕获或直接观察所提出的中间体。我们的具体目标2和3是研究Glu 57和Cys 84在B中的催化作用。通过诱变和化学拯救，以及Cys 84氧化的生物学相关性，对枯草杆菌LuxS进行了初步研究。最后，我们计划设计，合成和测试基于机制的抑制剂LuxS，特别是那些与活性位点锌离子相互作用。我们还将研究卤代呋喃酮，一组天然抗菌剂，如何灭活LuxS，特别是霍乱弧菌酶。此外，我们将测试LuxS抑制剂对群体感应、生物膜形成和相关细菌生理学的影响。