tailieunhanh - Báo cáo Y học: ER–resident chaperone interactions with recombinant antibodies in transgenic plants

In this study, we demonstrate that the folding and assembly of IgG in transgenic tobacco plants is orchestrated by BiP (binding protein), an endoplasmic reticulum resident chap-erone. Expression of BiP and calreticulin was examined in transgenic tobacco plants that express immunoglobulin chains, either singly or in combination to form IgG anti-body. BiP mRNA expression was lowest in wild-type nontransformed plants and those that expressed immuno-globulin light chain alone. | Eur. J. Biochem. 269 6042-6051 2002 FEBS 2002 doi ER-resident chaperone interactions with recombinant antibodies in transgenic plants James Nuttall1 Nicholas Vine2 Jane L. Hadlington1 Pascal Drake2 Lorenzo Frigerio1 t and Julian . Ma2 t 1 Department of Biological Sciences University of Warwick Coventry UK 2Unit of Immunology Department of Oral Medicine and Patholoyy vuv s Hospttal London UK In this study we demonstrate that the folding and assembly of IgG in transgenic tobacco plants is orchestrated by BiP binding protein an endoplasmic reticulum resident chaperone. Expression of BiP and calreticulin was examined in transgenic tobacco plants that express immunoglobulin chains either singly or in combination to form IgG antibody. BiP mRNA expression was lowest in wild-type nontransformed plants and those that expressed immunoglobulin light chain alone. Higher mRNA levels were detected in plants expressing fully assembled immunoglobulin light and heavy chains and the most abundant levels of RNA transcript were found in those plants that expressed immunoglobulin heavy chain alone. Estimation of total BiP demonstrated a similar pattern with the highest levels detected in plants expressing immunoglobulin heavy chain alone. Immunoprecipitation studies demonstrated that BiP was associated with immunoglobulin chains extracted from protoplast lysates but not from secreted fluids. Again most BiP was coprecipitated from plants expressing heavy chain only and those that produced full length IgG. The binding of BiP to Ig heavy chains was ATP-sensitive. Co-expression of heavy and light chain resulted in IgG assembly and displacement of BiP from the heavy chain as the amount of light chain increased. Although calreticulin mRNA and total protein levels varied in a similar manner to those of BiP in the transgenic plants there was no evidence for association between calreticulin and Ig chains by coimmunoprecipitation. The results indicate that .