Adenosine kinase of arabidopsis. Kinetic properties and gene expression
Adenosine kinase of arabidopsis. Kinetic properties and gene expression
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DOI：
10.1104/pp.124.4.1775
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发表时间：
2000-12-01
期刊：
影响因子：
7.4
通讯作者：
von Schwartzenberg, K
中科院分区：
文献类型：
--
作者：
Moffatt, BA;Wang, L;von Schwartzenberg, K
为了评估植物中腺苷回收的功能意义，从拟南芥中分离出编码两种腺苷激酶 (ADK) 亚型的 cDNA 和基因。 ADK1 和 ADK2 编码序列非常相似，分别具有 92% 和 89% 的氨基酸和核苷酸同一性。每个 cDNA 在大肠杆菌中过表达，并测定了每个亚型的催化活性。两种 ADK 均具有相似的催化特性，对腺苷的 K-m 和 V-max/K-m 分别为 0.3 至 0.5 muM 和 5.4 至 22 L min(-1) mg(-1) 蛋白质。细胞分裂素核苷 N-6(异戊烯基)腺苷的 K-m 和 V-max/K-m 分别为 3 至 5 muM 和 0.021 至 0.14 L min-1 mg-1 蛋白质，表明腺苷是两种 ADK 同工型的首选底物。在拟南芥植物中，两个 ADK 基因都是组成型表达，在茎和根中发现最高的稳态 mRNA 水平。 ADK1 转录水平普遍高于 ADK2。 ADK 酶活性反映了在叶、花和茎的免疫印迹中观察到的相对 ADK 蛋白水平，但对于根、长角果和干种子来说，情况较差。这些 ADK 的催化特性、组织积累和表达水平表明它们在拟南芥中腺苷酸的补救合成和甲基回收中发挥着关键的代谢作用。它们还可能有助于细胞分裂素的相互转化。
To assess the functional significance of adenosine salvage in plants, the cDNAs and genes encoding two isoforms of adenosine kinase (ADK) were isolated from Arabidopsis. The ADK1- and ADK2-coding sequences are very similar, sharing 92% and 89% amino acid and nucleotide identity, respectively. Each cDNA was overexpressed in Escherichia coli, and the catalytic activity of each isoform was determined. Both ADKs had similar catalytic properties with a K-m and V-max/K-m for adenosine of 0.3 to 0.5 muM and 5.4 to 22 L min(-1) mg(-1) protein, respectively. The K-m and V-max/K-m for the cytokinin riboside N-6(isopentenyl) adenosine are 3 to 5 muM and 0.021 to 0.14 L min-l mg-l protein, respectively, suggesting that adenosine is the preferred substrate for both ADK isoforms. In Arabidopsis plants, both ADK genes are expressed constitutively, with the highest steady-state mRNA levels being found in stem and root. ADK1 transcript levels were generally higher than those of ADK2. ADK enzyme activity reflected relative ADK protein levels seen in immunoblots for leaves, flowers, and stems but only poorly so for roots, siliques, and dry seeds. The catalytic properties, tissue accumulation, and expression levels of these ADKs suggest that they play a key metabolic role in the salvage synthesis of adenylates and methyl recycling in Arabidopsis. They may also contribute to cytokinin interconversion.