PROLINE METABOLISM/REGULATION OF PURINES/ENERGY TRANSFER

我们建议研究脯氨酸的可能作用 代谢系统在调节能量产生和调节中的作用 生物合成途径及其在细胞间隙调节中的作用 能量转移。基于对红细胞和 肝细胞，似乎是吡咯烷-5-羧酸还原酶 (聚合酶链式反应)活性可能调节磷酸戊糖途径活性 并导致从头开始的嘌呤核苷酸的刺激 生物合成。此外，聚合酶链式反应和Pro-Ox可能是 参与细胞质氧化还原电势向能量的转移- 利用细胞器。因为不寻常的高水平 在根瘤中的聚合酶链式反应和嘌呤生物合成活性 类杆菌中的ProOx(缺乏耗能的隔间 糖酵解能力)，以及极高的能量通量 从细胞质转移到类杆菌、甘氨酸结节 MAX是一个很好的模型系统，可以研究建议的 聚合酶链式反应和蛋白氧合酶在房室间能量传递中的作用 以及嘌呤核苷酸的生物合成。实验计划是 旨在考察这两个不同的但不是相互的 专属，聚合酶链式反应和蛋白氧合酶的功能(即对嘌呤的调节 核苷酸生物合成和细胞器间的能量转移) 上文建议的。具体目标是：1)本地化和 聚合酶链式反应，原氧合酶和其他酶的活性特征 脯氨酸代谢；2)确定二者之间的关系 聚合酶链式反应活性和嘌呤核苷酸合成；3)确定作用 在氧化还原电势的房室间转移中；以及 4)论证了聚合酶链式反应的生理意义和 完整组织中的ProOx。实验方法包括 聚合酶链式反应和原氧合酶亚细胞生化特性研究 分级和示踪研究。所取得的成果将 有助于我们理解细胞的一般机制 和亚细胞能量传递和代谢调节 真核生物。

We propose to examine the possible role of the proline metabolizing system in regulating energy-generating and biosynthetic pathways and in mediating intercompartmental energy transfer. Based on studies with erythrocytes and hepatocytes, it appears that pyrroline-5-carboxylate reductase (PCR) activity may modulate pentose phosphate pathway activity and result in the stimulation of de novo purine nucleotide biosynthesis. In addition PCR and proline oxidase (ProOx) may be involved in transfer of cytoplasmic redox potential into energy- utilizing organelles. Because of the uncommonly high levels of PCR and purine biosynthetic activity in nodules, the presence of ProOx in bacteroids (energy-consuming compartments lacking glycolytic capability), and the extremely high flux of energy transfer from the cytoplasm into bacteroids, nodules of Glycine max are an excellent model system to investigate the proposed roles of PCR and ProOx in intercompartmental energy transfer and purine nucleotide biosynthesis. The experimental plan is designed to examine the two different, but not mutually exclusive, functions of PCR and ProOx (i.e. regulation of purine nucleotide biosynthesis and energy transfer between organelles) proposed above. The specific objectives are: 1) to localize and characterize activities of PCR, ProOx, and other enzymes of proline metabolism; 2) to determine the relationship between PCR activity and purine nucleotide synthesis; 3) to define the role of proline in intercompartmental transfer of redox potential; and 4) to demonstrate the physiological significance of PCR and ProOx in intact tissues. The experimental approach involves biochemical characterization of PCR and ProOx, subcellular fractionation and tracer studies. The results obtained will contribute to our understanding of general mechanisms of cellular and subcellular energy transfer and metabolic regulation in eucaryotes.