Targeted protein accumulation promoted by autoassembly and its recovery from plant cells.
The expression of a fusion protein formed between the avian infectious bronchitis virus M protein and the bacterial enzyme beta-glucuronidase (GUS) in plants promotes the formation of new organization of the endoplasmic reticulum in tobacco plants. This unusual organization of the membranes, never present in nontransformed plants, has been explained by the oligomerization of the GUS domains of the IBVM-GUS fusion proteins. These specific organized membranes could have broad implications for biotechnology since their formation could be used as a mechanism for retaining and accumulating resident proteins in specific and discrete membrane compartments. In this study, we have shown that the unusual organization of native membranes due to overexpression of the IBVM-GUS fusion gene in tobacco transgenic plants and calli is present at higher levels in plant cell suspensions than in plant tissues. In these cell suspensions, IBVM-GUS protein was continuously synthesized and accumulated throughout the cell culture. An enrichment of the chimeric IBVM-GUS protein corresponding to a five-fold increase in the microsomal fractions was achieved and the GUS enzyme did not show any modification on enzyme kinetics. However, the GUS activity could be differentially distributed in the fractions eluted at different pH suggesting differences in the surface topography of histidine residues for this recombinant GUS.
Noizet
Mildred
M
UMR 6022 CNRS, Université de Technologie de Compiègne, Compiègne Cedex, France.
Harrabi
Faouzi
F
Vijayalakshmi
Mookambeswaran
M
Galbraith
David
D
Thomas
Daniel
D
Thomasset
Brigitte
B
D002478:Cells, Cultured; D018390:Gene Targeting / Q000379:methods*; D010940:Plant Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D014026:Nicotiana / Q000502:physiology*
D010940:Plant Proteins; D011993:Recombinant Fusion Proteins