Takase K, Hagiwara K
Department of Biotechnology, National Institute of Agrobiological Resources, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan.
J Biochem. 1998 Mar;123(3):440-4. doi: 10.1093/oxfordjournals.jbchem.a021956.
alphaLA-Lactalbumin (alphaLA), a major milk protein, is the regulatory subunit of lactose synthase. To assess the production of recombinant alphaLA in plants, the cDNAs for human alphaLA with or without its own signal sequence were introduced into tobacco plants under the control of the cauliflower mosaic virus 35S promoter. The gene integration and expression at the mRNA level were confirmed in several regenerated plants, while the expression at the protein level could be confirmed only in a transgenic tobacco transformed with the gene containing the signal sequence. The tobacco-expressed alphaLA migrated in SDS-PAGE with identical mobility to alphaLA prepared from human milk, indicating that the signal peptide of human alphaLA was correctly processed to yield a mature protein in tobacco plants. The expressed alphaLA (ca. 5 microg/g of fresh leaves) was found in the soluble fraction and eluted from a DEAE-Sepharose column in the same salt concentration range as the milk alphaLA. The partially purified tobacco-alphaLA was fully active in the synthesis of lactose when combined with galactosyltransferase. Thus, the transgenic tobacco produces a fully active mature alphaLA in a soluble form.
α-乳白蛋白（αLA）是一种主要的乳蛋白，是乳糖合酶的调节亚基。为了评估植物中重组αLA的产量，将带有或不带有自身信号序列的人αLA的cDNA在花椰菜花叶病毒35S启动子的控制下导入烟草植株。在几株再生植株中证实了基因整合以及mRNA水平的表达，而仅在转染了含有信号序列基因的转基因烟草中证实了蛋白质水平的表达。烟草表达的αLA在SDS-PAGE中的迁移率与人乳中制备的αLA相同，这表明人αLA的信号肽在烟草植株中被正确加工产生了成熟蛋白。表达的αLA（约5微克/克鲜叶）存在于可溶部分，并且从DEAE-琼脂糖柱上洗脱的盐浓度范围与乳αLA相同。部分纯化的烟草αLA与半乳糖基转移酶结合时，在乳糖合成中具有完全活性。因此，转基因烟草产生了可溶形式的具有完全活性的成熟αLA。
