Rodaway S J, Kende H
Michigan State University Energy Research and Development Administration, Plant Research Laboratory Michigan State University, East Lansing, Michigan 48824.
Plant Physiol. 1978 Jan;61(1):1-6. doi: 10.1104/pp.61.1.1.
alpha-Amylase has been purified from de-embryonated seeds of barley (Hordeum vulgare L. cv. Betzes) which have been incubated on 10(-6)m gibberellic acid (GA(3)) following 3 days of imbibition in buffer. Incubation of the half-seeds in up to 10(-2)m 5-fluorouracil (5-FU) during the entire incubation period, including imbibition, had no effect on any of the following characteristics of purified alpha-amylase: thermal stability in the absence of calcium, molecular weight of the enzyme, isozyme composition, specific activity, or the amount of alpha-amylase synthesized by the aleurone tissue. The synthesis of rRNA and tRNA was strongly inhibited by 5-FU, indicating that the analog had entered the aleurone cells. These results are not in agreement with those of Carlson (Nature New Biology 237: 39-41 [1972]) who found that treatment of barley aleurone with 10(-4)m 5-FU prior to the addition of GA(3) resulted in decreased thermal stability of GA(3)-induced alpha-amylase and who interpreted this as evidence that the mRNA for alpha-amylase was synthesized during the imbibition of the aleurone tissue and independently of gibberellin action. Results of the present experiments indicate that the thermal stability of highly purified alpha-amylase is not altered by treatment of barley half-seeds with 5-FU, and that 5-FU cannot be used as a probe to examine the timing of alpha-amylase mRNA synthesis.
已从大麦（Hordeum vulgare L. cv. Betzes）的去胚种子中纯化出α-淀粉酶。这些种子在缓冲液中吸胀3天后，在10⁻⁶m赤霉素（GA₃）中孵育。在整个孵育期（包括吸胀期），将半粒种子在高达10⁻²m的5-氟尿嘧啶（5-FU）中孵育，对纯化的α-淀粉酶的以下任何特性均无影响：无钙时的热稳定性、酶的分子量、同工酶组成、比活性或糊粉层组织合成的α-淀粉酶量。5-FU强烈抑制rRNA和tRNA的合成，表明该类似物已进入糊粉层细胞。这些结果与卡尔森（《自然新生物学》237: 39 - 41 [1972]）的结果不一致，卡尔森发现，在添加GA₃之前用10⁻⁴m 5-FU处理大麦糊粉层会导致GA₃诱导的α-淀粉酶热稳定性降低，他将此解释为α-淀粉酶的mRNA是在糊粉层组织吸胀期间合成且独立于赤霉素作用的证据。本实验结果表明，用5-FU处理大麦半粒种子不会改变高度纯化的α-淀粉酶的热稳定性，并且5-FU不能用作检测α-淀粉酶mRNA合成时间的探针。
