Amidophosphoribosyltransferase limits the rate of cell growth-linked de novo purine biosynthesis in the presence of constant capacity of salvage purine biosynthesis
Amidophosphoribosyltransferase limits the rate of cell growth-linked de novo purine biosynthesis in the presence of constant capacity of salvage purine biosynthesis
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DOI：
10.1074/jbc.272.28.17719
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发表时间：
1997-07-11
影响因子：
4.8
通讯作者：
Itakura, M
中科院分区：
文献类型：
--
作者：
Yamaoka, T;Kondo, M;Itakura, M
在动物细胞生长过程中，控制嘌呤核苷酸生物合成从头合成和挽救途径的相对通量速率的因素尚不完全清楚，无法检查每条途径对细胞的相对作用。将CHO K1细胞系(野生型中国仓鼠卵巢成纤维细胞系)、CHO ADE(-)A细胞系(缺失从头途径的限速酶)和CHO ADE(-)A细胞系(转染人ATase基因(-)A+hATase)，使其AT酶活性提高30%~350%，分别在富嘌呤和去嘌呤的培养液中培养，研究AT酶和次黄嘌呤磷酸核糖转移酶的代谢速率和回收途径三种细胞系在不同培养条件下的生长速度以及次黄嘌呤对大鼠肝脏从头途径代谢率的影响，我们得出以下结论：1)在(-)A+hATase转染体中，ATase活性限制了与细胞生长速度密切相关的从头途径的速率；2)嘌呤核苷酸的优先合成途径是抢救途径，只要次黄嘌呤是嘌呤挽救的最主要来源，这一点在体内大鼠肝脏中得到证实，挽救途径的优先使用导致了从头合成所需的能量支出的节省。3)次黄嘌呤磷酸核糖转移酶活性恒定的从头途径的调节能力(约200%)大于挽救途径(约20%)。
Factors controlling relative flux rates of the de novo and salvage pathways of purine nucleotide biosynthesis during animal cell growth are not fully understood, To examine the relative role of each pathway for cell. growth, three cell lines including CHO K1 (a wild-type Chinese hamster ovary fibroblast cell line), CHO ade (-)A (an auxotrophic cell line deficient of amidophosphoribosyltransferase (ATase), a presumed rate-limiting enzyme of the de novo pathway), and CHO ade (-)A transfected with human ATase cDNA ((-)A+hATase) resulting in 30-350% of the ATase activity of CHO K1, were cultured in purine-rich or purine-free media, Based on the enzyme activities of ATase and hypoxanthine phosphoribosyltransferase, the metabolic rate of the de novo and salvage pathways, the rate of cell growth (growth rate) in three cell lines under various culture conditions, and the effect of hypoxanthine infusion on the metabolic rate of the de novo pathway in rat liver, we concluded the following, 1) In (-)A+hATase transfectants, ATase activity limits the rate of the de novo pathway, which is closely linked with the growth rate, 2) Purine nucleotides are synthesized preferentially by the salvage path way as long as hypoxanthine, the most essential source of purine salvage, can be utilized, which was confirmed in rat liver in vivo by hypoxanthine infusion, The preferential usage of the salvage pathway results in sparing the energy expenditure required for de novo synthesis, 3) The regulatory capacity of the de novo pathway (about 200%) was larger than that of the salvage pathway (about 20%) with constant hypoxanthine phosphoribosyltransferase activity.