H2S production and virulence of Treponema denticola

齿垢密螺旋体的 H2S 产生和毒力

• 批准号: 7004557
• 负责人: LIANRUI CHU
• 金额: $ 20.32万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7004557
• 关键词: Treponema; Treponema infection; affinity chromatography; bacterial proteins; clinical research; cysteine; enzyme activity; enzyme inhibitors; gene induction /repression; glutathione; high performance liquid chromatography; human subject; hydrogen sulfide; immunoelectron microscopy; laboratory mouse; laboratory rabbit; mass spectrometry; molecular cloning; nucleic acid sequence; oral bacteria; pathologic process; periodontitis; periodontium disorder; polymerase chain reaction; protein glutamine gamma glutamyltransferase; virulence; western blottings

DESCRIPTION: This application is an extension of current research examining the association between H2S production and virulence of Treponema denticola. T. denticola has been identified as an important member of a consortium of microorganisms as etiologic in the initiation and progression of periodontal diseases. In addition, the existence of volatile sulfur compounds produced at destructive sites is a characteristic feature of periodontal diseases, with H2S as a major compound in this family. Nevertheless, the metabolic pathways used to produce H2S are not well understood. Previous studies suggest that glutathione present in host cells can be a substrate for H2S production. Recently, we have shown that T. denticola has the capacity to utilize glutathione as a substrate to produce high levels of H2S. We have identified three enzymes that are required for the successful metabolism of glutathione: gamma-glutamyltransferase (GGT), cysteinylglycinase (CGase), and cystalysin. GGT converts glutathione into Cys-Gly and glutamic acid; CGase catalyze Cys-Gly to Cys and glycine; and cystalysin digests L-cysteine into H2S, ammonia, and pyruvate. We have also demonstrated that the addition of cystalysin and L-cysteine resulted in apoptosis of HGF and PDL cells and that glutathione was essential for lesion formation by T. denticola in an animal model. Based on these and other studies, three Specific Aims are proposed using biochemical, molecular genetic, and cell biologic studies to address the hypothesis that these three metabolic enzymes play a key role in T denticola pathogenesis. Specific Aim 1: To molecularly characterize the genes and proteins involved in converting glutathione to H2S. Specific Aim 2: To genetically characterize, by gene inactivation, the enzyme pathway of T. denticola that produces H2S from glutathione. Specific Aim 3: To measure the effects of the H2S/NH3 producing pathway on T. denticola virulence in vitro and in vivo. This application is designed to provide both seminal and critical information about the enzyme pathway of T. denticola to produce H2S from glutathione. The outcomes will elucidate the mechanisms of action that each member of the enzyme pathway plays in the virulence capacity of T. denticola.

描述：这个应用程序是目前研究的H2S生产和密螺旋体毒力之间的关系的延伸。牙周菌已被确定为牙周病发生和发展的病原学微生物联盟的重要成员。此外，在破坏性部位产生的挥发性硫化合物的存在是牙周病的一个特征，其中H2S是该家族的主要化合物。然而，用于产生H2S的代谢途径尚不清楚。先前的研究表明，宿主细胞中的谷胱甘肽可能是H2S生产的底物。最近，我们已经证明T. denticola有能力利用谷胱甘肽作为底物产生高水平的H2S。我们已经确定了谷胱甘肽成功代谢所必需的三种酶：γ -谷氨酰转移酶（GGT）、半胱氨酸甘氨酸酶（gase）和囊溶素。GGT将谷胱甘肽转化为ys- gly和谷氨酸；case催化Cys- gly生成Cys和甘氨酸；胱氨酸将l -半胱氨酸分解成H2S、氨和丙酮酸。我们还证明，在动物模型中，添加囊溶素和l -半胱氨酸可导致HGF和PDL细胞凋亡，谷胱甘肽对齿状鼠的病变形成至关重要。在此基础上，本文提出了三个具体目标，通过生物化学、分子遗传学和细胞生物学的研究来解决这三种代谢酶在树突线虫发病机制中起关键作用的假设。具体目标1：分子表征参与将谷胱甘肽转化为H2S的基因和蛋白质。具体目标2：通过基因失活，从基因上表征T. denticola从谷胱甘肽中产生H2S的酶途径。专项目的3：测定H2S/NH3产生途径对牙鼠体外和体内毒力的影响。该应用程序旨在提供关于T. denticola从谷胱甘肽产生H2S的酶途径的开创性和关键信息。结果将阐明作用机制，酶途径的每个成员在齿状霉的毒力能力发挥作用。