HEPATIC GLUTAMINASE

肝谷氨酰胺酶

基本信息

批准号：
3236143
负责人：
MALCOLM WATFORD
金额：
$ 11.23万
依托单位：
RUTGERS, THE STATE UNIV OF N.J.
依托单位国家：
美国
项目类别：
财政年份：
1989
资助国家：
美国
起止时间：
1989-07-01 至 1992-12-31
项目状态：
已结题

项目摘要

Glutamine plays an important biosynthetic role in all mammalian cells. Glutamine is used by the liver during diabetes, the kidney during metabolic acidosis and the mammary gland during lactation, it is also a major respiratory fuel for many rapidly dividing cells including cancer cells. The major enzymes of glutamine catabolism in mammalian tissues are two isozymes of phosphate activated glutaminase. The liver type is found exclusively in adult liver while the other enzyme (kidney-type) is found in all other glutamine utilizing tissues, including fetal liver and hepatoma cells. The aim of the work described is to characterize and understand the role and regulation of phosphate activated glutaminase in rat liver. This work stems from the observation that hepatic glutaminase activity is increased 4-fold during streptozotocin diabetes. The enzyme has been purified, specific antibodies raised against it and used to identify a putative cDNA. The identify of the cDNA will be confirmed by comparison of the DNA and amino acid sequences, hybrid selection of mRNA and analysis of fusion peptide. The cDNA will be used to determine the relative abundance of mRNA and the rate of transcription hepatic glutaminase gene in liver from control and diabetic rats. Extracellular stimuli involved in the regulation of hepatic glutaminase gene expression will be identified using isolated hepatocytes and hepatoma cells. The cDNA will be used to identify genomic sequences encoding liver-type glutaminase and other glutamine utilizing enzymes. The distribution of liver- and kidney-type glutaminase (activity, protein and mRNA) will be determined in different hepatoma cells and in rat liver at different stages of development. Chromatin structure will be analyzed in cells showing differential expression of the two genes. These systems offer a unique model for the study of tissue-specific, hormonal and developmental regulation of glutaminase gene expression. The results will provide valuable insights into why there are two enzymes and what their functions are in the different cell types.

谷氨酰胺在所有哺乳动物细胞中起着重要的生物合成作用。谷氨酰胺在糖尿病期间由肝脏使用，在糖尿病期间由肾脏使用代谢性酸中毒和哺乳期间的乳腺，它也是一个许多快速分裂细胞（包括癌症）的主要呼吸燃料细胞。哺乳动物组织中谷氨酰胺分解代谢的主要酶是磷酸激活谷氨酰胺酶的两种同工酶。肝脏类型是仅在成人肝脏中发现，而另一种酶（肾型）是存在于所有其他利用谷氨酰胺的组织中，包括胎儿肝脏和肝癌细胞。所描述的工作的目的是表征和了解磷酸激活谷氨酰胺酶的作用和调节大鼠肝脏。这项工作源于肝谷氨酰胺酶活性的观察链脲佐菌素糖尿病期间增加 4 倍。该酶已针对它产生的纯化的特异性抗体并用于识别推定的cDNA。通过比对确认cDNA的鉴定 DNA 和氨基酸序列的分析、mRNA 和 mRNA 的杂交选择融合肽的分析。 cDNA 将用于确定 mRNA相对丰度和肝脏转录率对照组和糖尿病大鼠肝脏中的谷氨酰胺酶基因。细胞外参与肝谷氨酰胺酶基因表达调节的刺激将使用分离的肝细胞和肝癌细胞进行鉴定。该 cDNA 将用于鉴定编码肝型的基因组序列谷氨酰胺酶和其他利用谷氨酰胺的酶。的分布肝型和肾型谷氨酰胺酶（活性、蛋白质和 mRNA）将在不同的肝癌细胞和不同的大鼠肝脏中测定发展阶段。将在细胞中分析染色质结构显示两个基因的差异表达。这些系统提供了用于研究组织特异性、激素和发育的独特模型谷氨酰胺酶基因表达的调控。结果将提供关于为什么存在两种酶及其功能的宝贵见解位于不同的细胞类型中。