Role of BPHL/homocysteine thiolactonase in hyperhomocysteinemia and disease

BPHL/同型半胱氨酸硫内酯酶在高同型半胱氨酸血症和疾病中的作用

• 批准号: 9324453
• 负责人: Judit Marsillach Lopez
• 金额: $ 34.78万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-06 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9324453
• 关键词: Address; Albumins; Alzheimer' s Disease; Amino Acids; Apolipoprotein A-I; Architecture; Arylesterase; Atherosclerosis; Autoimmune Diseases; Binding; Biological; Biological Markers; Blood; Brain; Cardiovascular Diseases; Clinical; Clinical Research; Congenital Abnormality; Data; Detection; Development; Diet; Disease; Disease Marker; Distress; Dose; Drug Metabolic Detoxication; Enzymes; Foundations; Future; Gene Expression; Genetic; Homocysteine; Homocystine; Human; Hydrolysis; Hyperhomocysteinemia; Immunohistochemistry; Inflammation; Injection of therapeutic agent; Kidney; Kidney Failure; Knock-out; Knockout Mice; Laboratories; Lead; Link; Liver; Lysine; Mass Spectrum Analysis; Mental disorders; Methionine; Methionine-tRNA Ligase; Methods; Modification; Monitor; Mus; Nerve Degeneration; Organ; Pathology; Physiological; Plasma; Plasma Proteins; Play; Population; Prodrugs; Proteins; Protocols documentation; RNA; Recombinants; Research; Risk Factors; Role; Saline; Seizures; Study models; Tissues; Toxic effect; Transgenic Mice; Urine; Valganciclovir; Vitamin B Complex; Vitamin B Deficiency; Wild Type Mouse; Work; adduct; aortic arch; base; biphenyl hydrolase-related protein; bleomycin hydrolase; disorder risk; enzyme activity; experience; feeding; histological stains; improved; in vivo; insight; loss of function; nervous system disorder; novel; prevent; protein biomarkers; valacyclovir

PROJECT SUMMARY/ABSTRACT Elevated levels of the amino acid homocysteine (Hcy) are a risk factor for a wide variety of disorders, from cardiovascular disease to neurological and autoimmune disorders. The mechanisms underlying these effects are not well understood. High plasma levels of Hcy lead to high levels of homocysteine thiolactone (HCTL), a toxic metabolite of Hcy. HCTL has the ability to interact with and modify proteins, resulting in protein inactivation and loss of function. The biological significance and consequence of these modifications is not well known, although they may trigger the development of the diseases noted above. We have recently characterized a very efficient homocysteine thiolactonase activity of the enzyme biphenyl hydrolase-like protein (BPHL), previously studied for its role in bioactivating the prodrugs valacyclovir and valganciclovir. Although two other enzymes with homocysteine thiolactonase activity, paraoxonase-1 (PON1) and bleomycin hydrolase (Blmh), had previously been described, BPHL has a much higher catalytic efficiency for HCTL, indicating a more significant physiological role for BPHL in detoxifying HCTL. The proposed research aims to establish the extent to which BPHL protects against HCTL toxicity and the development of diseases associated with high Hcy/HCTL levels. In addition, we will demonstrate that in conditions of hyperhomocysteinemia, plasma proteins that are modified by HCTL can serve as biomarkers of HCTL-associated disease risk. The following proposed specific aims make use of the commercially-available BPHL knockout (BPHL-/-) mouse and our experience in identifying protein adducts by mass spectrometry (MS): Aim 1 will examine the effects of a [non-toxic (sub-Aim 1a) or toxic (sub-Aim 1b)] dose injection of HCTL on BPHL-/- and BPHL+/+ (wild-type) mice compared with controls injected with saline. The plasma proteins albumin and apolipoprotein A-I will be analyzed for HCTL adducts on lysine residues, and the urine for higher levels of secreted HCTL using MS analyses. Aim 2 will examine the ability of BPHL-/- mice to modulate the effects of hyperhomocysteinemia generated by diets high in methionine [with or without deficiency in B vitamins]. We will examine mouse tissues for [early] signs of pathology, together with effects on gene expression (sub-Aim 2a). We will also identify HCTL adducts on the same proteins monitored in Aim 1 (sub-Aim 2b). These studies elucidate a novel physiological function of BPHL. Since the physiological significance of BPHL was unknown prior to our recent work, the results obtained from the proposed research have the potential to define the role of BPHL as that of a physiologically significant, protective homocysteine thiolactonase. We will also develop improved MS-based methods for monitoring HCTL adducts. The results obtained from this proposal will increase our understanding of the link between hyperhomocysteinemia and development of disease, and provide a basis for examining the effects of genetic variability in BPHL as a risk factor for HCTL- associated diseases.

项目总结/摘要 氨基酸同型半胱氨酸（Hcy）水平升高是多种疾病的危险因素， 从心血管疾病到神经系统和自身免疫性疾病。这些影响的机制 并没有得到很好的理解。高血浆同型半胱氨酸水平导致高水平的同型半胱氨酸硫内酯（HCTL）， 同型半胱氨酸的毒性代谢产物。HCTL具有与蛋白质相互作用并修饰蛋白质的能力，从而产生蛋白质。 失活和功能丧失。这些修饰的生物学意义和后果尚不清楚 虽然它们可能会引发上述疾病的发展。我们最近 其特征在于一种具有非常高效的同型半胱氨酸硫内酯酶活性的酶联苯水解酶样蛋白 （BPHL），先前研究其在生物活化前药伐昔洛韦和缬更昔洛韦中的作用。虽然 另外两种具有同型半胱氨酸硫内酯酶活性的酶，对氧磷酶-1（PON 1）和博来霉素水解酶 （Blmh），BPHL对HCTL具有高得多的催化效率，表明BPHL对HCTL具有高得多的催化效率。 BPHL对HCTL的解毒作用更为显著。 该研究旨在确定BPHL对HCTL毒性的保护程度以及BPHL对HCTL毒性的保护程度。 与高Hcy/HCTL水平相关的疾病的发展。此外，我们将证明，在 在高同型半胱氨酸血症的情况下，被HCTL修饰的血浆蛋白可以作为高同型半胱氨酸血症的生物标志物。 HCT相关疾病风险。以下提出的具体目标利用了商业上可获得的 BPHL敲除（BPHL-/-）小鼠和我们通过质谱（MS）鉴定蛋白加合物的经验： 目标1将检查[无毒（子目标1a）或毒性（子目标1b）]剂量注射HCTL对 BPHL-/-和BPHL+/+（野生型）小鼠与注射生理盐水的对照相比。血浆蛋白白蛋白 和载脂蛋白A-I将分析赖氨酸残基上的HCTL加合物，并分析尿液中较高水平的 使用MS分析分泌的HCTL。目的2将检查BPHL-/-小鼠调节以下作用的能力： 由高蛋氨酸饮食[有或没有B族维生素缺乏]引起的高同型半胱氨酸血症。我们将 检查小鼠组织的病理学[早期]迹象，以及对基因表达的影响（子目标2a）。 我们还将鉴定目标1中监测的相同蛋白质上的HCTL加合物（子目标2b）。 这些研究阐明了BPHL的一种新的生理功能。由于BPHL的生理意义 在我们最近的工作之前是未知的，从拟议的研究中获得的结果有可能 将BPHL的作用定义为生理学上显著的、保护性的同型半胱氨酸硫内酯酶。我们将 还开发了用于监测HCTL加合物的改进的基于MS的方法。由此获得的结果 该提案将增加我们对高同型半胱氨酸血症与发展之间联系的理解。 疾病，并提供了一个基础，检查遗传变异的影响，在BPHL作为一个危险因素的HCTL- 相关疾病。