Grantz A A, Brummell D A, Bennett A B
Mann Laboratory, Department of Vegetable Crops, University of California, Davis 95616, USA.
Plant Physiol. 1995 May;108(1):411-8. doi: 10.1104/pp.108.1.411.
A cDNA clone encoding ascorbate free radical (AFR) reductase (EC 1.6.5.4) was isolated from tomato (Lycopersicon esculentum Mill.) and its mRNA levels were analyzed. The cDNA encoded a deduced protein of 433 amino acids and possessed amino acid domains characteristic of flavin adenine dinucleotide- and NAD(P)H-binding proteins but did not possess typical eukaryotic targeting sequences, suggesting that it encodes a cytosolic form of AFR reductase. Low-stringency genomic DNA gel blot analysis indicated that a single nuclear gene encoded this enzyme. Total ascorbate contents were greatest in leaves, with decreasing amounts in stems and roots and relatively constant levels in all stages of fruit. AFR reductase activity was inversely correlated with total ascorbate content, whereas the relative abundance of AFR reductase mRNA was directly correlated with enzyme activity in tissues examined. AFR reductase mRNA abundance increased dramatically in response to wounding, a treatment that is known to also induce ascorbate-dependent prolyl hydroxylation required for the accumulation of hydroxyproline-rich glycoproteins. In addition, AFR reductase may contribute to maintaining levels of ascorbic acid for protection against wound-induced free radical-mediated damage. Collectively, the results suggest that AFR reductase activity is regulated at the level of mRNA abundance by low ascorbate contents or by factors that promote ascorbate utilization.
从番茄（Lycopersicon esculentum Mill.）中分离出一个编码抗坏血酸自由基（AFR）还原酶（EC 1.6.5.4）的cDNA克隆，并对其mRNA水平进行了分析。该cDNA编码一个由433个氨基酸组成的推导蛋白，具有黄素腺嘌呤二核苷酸和NAD（P）H结合蛋白的特征性氨基酸结构域，但不具有典型的真核靶向序列，这表明它编码一种胞质形式的AFR还原酶。低严谨度基因组DNA凝胶印迹分析表明，该酶由单个核基因编码。总抗坏血酸含量在叶片中最高，在茎和根中含量逐渐降低，在果实的所有阶段含量相对恒定。AFR还原酶活性与总抗坏血酸含量呈负相关，而AFR还原酶mRNA的相对丰度与所检测组织中的酶活性呈正相关。AFR还原酶mRNA丰度在受伤后显著增加，已知这种处理也会诱导富含羟脯氨酸糖蛋白积累所需的抗坏血酸依赖性脯氨酰羟化。此外，AFR还原酶可能有助于维持抗坏血酸水平，以保护免受伤口诱导的自由基介导的损伤。总体而言，结果表明AFR还原酶活性在mRNA丰度水平上受低抗坏血酸含量或促进抗坏血酸利用的因素调节。
