Koc Ahmet, Mathews Christopher K, Wheeler Linda J, Gross Michael K, Merrill Gary F
Department of Biochemistry and Biophysics, Oregon State University, Corvallis, Oregon 97331, USA.
J Biol Chem. 2006 Jun 2;281(22):15058-63. doi: 10.1074/jbc.M601968200. Epub 2006 Mar 29.
Thioredoxin was initially identified by its ability to serve as an electron donor for ribonucleotide reductase in vitro. Whether it serves a similar function in vivo is unclear. In Saccharomyces cerevisiae, it was previously shown that Deltatrx1 Deltatrx2 mutants lacking the two genes for cytosolic thioredoxin have a slower growth rate because of a longer S phase, but the basis for S phase elongation was not identified. The hypothesis that S phase protraction was due to inefficient dNTP synthesis was investigated by measuring dNTP levels in asynchronous and synchronized wild-type and Deltatrx1 Deltatrx2 yeast. In contrast to wild-type cells, Deltatrx1 Deltatrx2 cells were unable to accumulate or maintain high levels of dNTPs when alpha-factor- or cdc15-arrested cells were allowed to reenter the cell cycle. At 80 min after release, when the fraction of cells in S phase was maximal, the dNTP pools in Deltatrx1 Deltatrx2 cells were 60% that of wild-type cells. The data suggest that, in the absence of thioredoxin, cells cannot support the high rate of dNTP synthesis required for efficient DNA synthesis during S phase. The results constitute in vivo evidence for thioredoxin being a physiologically relevant electron donor for ribonucleotide reductase during DNA precursor synthesis.
硫氧还蛋白最初是因其在体外作为核糖核苷酸还原酶的电子供体的能力而被鉴定出来的。它在体内是否发挥类似功能尚不清楚。在酿酒酵母中，先前的研究表明，缺乏胞质硫氧还蛋白两个基因的Δtrx1Δtrx2突变体由于S期延长而生长速率较慢，但S期延长的原因尚未确定。通过测量异步和同步的野生型及Δtrx1Δtrx2酵母中的脱氧核苷酸三磷酸（dNTP）水平，研究了S期延长是由于dNTP合成效率低下这一假说。与野生型细胞不同，当α因子或cdc15阻滞的细胞被允许重新进入细胞周期时，Δtrx1Δtrx2细胞无法积累或维持高水平的dNTP。在释放后80分钟，当处于S期的细胞比例最大时，Δtrx1Δtrx2细胞中的dNTP池是野生型细胞的60%。数据表明，在缺乏硫氧还蛋白的情况下，细胞无法支持S期高效DNA合成所需的高dNTP合成速率。这些结果构成了体内证据，证明硫氧还蛋白在DNA前体合成过程中是核糖核苷酸还原酶的生理相关电子供体。
