Metabolic Pathways and Defects in Fructose Metabolism

果糖代谢的代谢途径和缺陷

基本信息

批准号：
7850237
负责人：
Dean R. TOLAN
金额：
$ 1.63万
依托单位：
BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-07-01 至 2010-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7850237
关键词：
African American Aldehyde-Lyases Aldolase B Algorithms Alleles American Animal Model Animals Assimilations Bacteria Binding Biochemical Biochemical Genetics Bioinformatics Biological Assay Blood specimen Clinical Code Complement Complement component C1s Complex Corticosterone Crystallography DNA Microarray Chip DNA Sequence DNA Sequence Determination Data Databases Defect Development Diabetes Mellitus Diagnosis Diagnostic Diagnostic Procedure Diet Digestion Disease Drug Design Enzymes Ethnic group Exons Expressed Sequence Tags Family Fasting Fatty Acids Fructokinases Fructose GLUT2 gene Gene Expression Gene Targeting Genes Genetic Structures Genetic Transcription Genotype Glucose Goals Health Heart Diseases Hereditary Disease Hereditary fructose intolerance syndrome Hexose Transporter Hispanic Americans Hispanics Human Immunohistochemistry In Situ Hybridization Inborn Genetic Diseases Incubated Individual Ingestion Insulin Investigation Isoenzymes Ketohexokinase Kidney Knock-in Mouse Knock-out Knockout Mice Knowledge Laboratories Libraries Ligands Liver Messenger RNA Metabolic Metabolic Pathway Metabolism Modeling Molecular Molecular Models Molecular Profiling Mus Mutation Obesity Oligonucleotides Operative Surgical Procedures Other Genetics Pathology Pathway interactions Pattern Pharmaceutical Preparations Phenotype Phosphotransferases Physiological Physiology Play Population Procedures RNA Radioactive Recovery Resolution Reverse Transcriptase Polymerase Chain Reaction Role Screening procedure Site Slice Small Intestines Structure Techniques Technology Temperature Testing Therapeutic Therapeutic Agents Tissues Triglycerides Trioses Validation X-Ray Crystallography alternative treatment base cell type chemical genetics combinatorial chemistry design diabetic dimer embryonic stem cell fructose-1,6-diphosphate fructose-1-phosphate fructose-6-phosphate hexokinase high throughput screening molecular modeling mouse model mutant novel oxidation recombinase research study restriction enzyme segregation small molecule small molecule libraries sugar therapeutic development tool triokinase vector

项目摘要

DESCRIPTION (provided by applicant): Long-term ingestion has effects on diabetes and obesity. The most drastic and common genetic disorder of fructose metabolism is hereditary fructose intolerance (HFI). Lack of knowledge about sites of fructose metabolism and about genotype-phenotype relationships still exists for this disease and reflects the incomplete understanding of normal fructose metabolism. Answers to two major questions will provide new information. First, other than liver and kidney, what other tissues play a role in fructose metabolism? Second, can small molecules be found that stabilize the major defective enzyme in HFI, that harboring an A149P substitution (AP-aldolase)? The proposed investigations will, 1) define sites for fructose assimilation and utilization using a combination of bioinformatics and molecular approaches, 2) determine a high-resolution structure of AP-aldolase, and use it to find stabilizing small-molecule ligands by both structure-based ligand design (SBLD) and high-throughput screening of chemical libraries, 3) create animal models for HFI using gene-targeting techniques, and 4) identify HFI mutations in the diverse US population, in particular Hispanic, African-American, and other ethnic groups that have not been well characterized, and correlate these findings to any specific phenotypes in these ethnic groups. The large database of expressed sequence tags (dbEST) will be analyzed for overlapping expression profiles of the GLUT5, GLUT2, ketohexokinase, aldolase, hexokinase, and triose kinase in both mouse and humans to predict alternative sites of fructose metabolism. Verification and characterization of these global predictions will be done by quantitative reverse-transcriptase polymerase chain reaction (Q-PCR), RNA in situ hybridization (RISH), and identification of metabolic intermediates during the oxidation of radioactive fructose. For the second hypothesis, a high-resolution structure of AP-aldolase will be determined by macromolecular X-ray crystallography. Screening large libraries of small molecules (produced by combinatorial chemistry) using a thermal-stability assay of AP-aldolase in conjunction with SBLD will identify small molecules that restore enzyme function. Should the gene-targeted animal model mimic the human HFI pathology, the metabolic profiles and sites of fructose metabolism will be determined. Should the animal model be asymptomatic, differences in the ability to metabolize fructose will be compared to previously determined sites and pathways for fructose metabolism in normal mice and humans. Lastly, blood samples from African-American and Hispanic-American HFI subjects will be used for identification of gene defects by direct DNA sequencing, thus offering a reliable non-invasive diagnostic method to these Americans.

描述（由申请人提供）：长期摄入对糖尿病和肥胖症有影响。果糖代谢的最激烈，最常见的遗传疾病是遗传性果糖不耐症（HFI）。缺乏对果糖代谢部位以及对这种疾病的基因型 - 表型关系的知识，并反映了对正常果糖代谢的不完全理解。两个主要问题的答案将提供新的信息。首先，除了肝脏和肾脏，其他哪些组织在果糖代谢中起作用？其次，可以发现稳定HFI中主要有缺陷酶的小分子，该酶具有A149p替代（AP-钙化酶）？拟议的研究将1）使用生物信息学和分子方法的组合定义用于果糖同化和利用的位点，2）确定AP-溶解酶的高分辨率结构，并使用它来稳定地稳定基于结构的小分子配体，并使用基于结构的工具（SBLD）筛选型号的型号的小分子配体，以及3. sefterii creation sexing creation Inalies（SBLD）的3个型号。靶向基因的技术，以及4）确定美国多样化人群的HFI突变，尤其是西班牙裔，非裔美国人和其他尚未得到充分特征的族裔，并将这些发现与这些种族中的任何特定表型相关联。将分析表达序列标签的大数据库（DBEST），以分析小鼠和人类中GLUT2，Glut2，Ketohecokinase，aldolase，Hexokinase，Hexokinase和Triose激酶的重叠表达谱，以预测果糖代谢的交替位点。这些全局预测的验证和表征将通过定量反向转录酶聚合酶链反应（Q-PCR），RNA原位杂交（RISH）以及在放射性果糖氧化过程中鉴定代谢中间体。对于第二个假设，将通过大分子X射线晶体学确定AP-醛固酶的高分辨率结构。使用与SBLD结合的热溶解酶的热稳定性测定法筛选小分子（由组合化学产生）的大型文库（由组合化学产生）将确定恢复酶功能的小分子。如果以基因为靶向的动物模型模仿人类HFI病理，则将确定果糖代谢的代谢特征和部位。如果动物模型是无症状的，则将代谢果糖的能力差异与正常小鼠和人类果糖代谢的先前确定的位点和途径进行比较。最后，来自非裔美国人和西班牙裔美国人HFI受试者的血液样本将用于通过直接DNA测序来识别基因缺陷，从而向这些美国人提供可靠的非侵入性诊断方法。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

A six-month-old infant with liver steatosis.

一名六个月大的婴儿患有肝脏脂肪变性。

DOI：
10.1016/j.jpeds.2003.11.037
发表时间：
2004
期刊：
The Journal of pediatrics
影响因子：
0
作者：
Stormon,MichaelO;Cutz,Ernest;Furuya,Katryn;Bedford,Melanie;Yerkes,Laura;Tolan,DeanR;Feigenbaum,Annette
通讯作者：
Feigenbaum,Annette

Increased prevalence of mutant null alleles that cause hereditary fructose intolerance in the American population.

DOI：
10.1007/s10545-009-9008-7
发表时间：
2010-02
期刊：
JOURNAL OF INHERITED METABOLIC DISEASE
影响因子：
4.2
作者：
Coffee, Erin M.;Yerkes, Laura;Ewen, Elizabeth P.;Zee, Tiffany;Tolan, Dean R.
通讯作者：
Tolan, Dean R.

Specific regions of the brain are capable of fructose metabolism.