Expression of human alpha-lactalbumin in transgenic tobacco.
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.
Takase
K
K
Department of Biotechnology, National Institute of Agrobiological Resources, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan. ktakase@abr.affrc.go.jp
Hagiwara
K
K
D003001:Cloning, Molecular; D006801:Humans; D007768:Lactalbumin / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics*; D010947:Plants, Toxic; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
D011994:Recombinant Proteins; D007768:Lactalbumin