ISSN:
1573-9368
Keywords:
intracellular targeting
;
Nicotiana tabacum
;
prenyl transferase
;
signal peptide
;
ubiA
;
ubiquinone
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract The ubiA gene from E. coli codes for 4-hydroxybenzoate: polyprenyldiphosphate 3-polyprenyltransferase, an integral membrane protein involved in ubiquinone biosynthesis. This prokaryotic membrane protein was stably expressed in tobacco using Agrobacterium tumefaciens-mediated transformation. Transgenic lines containing a direct fusion of the ubiA structural gene to a 35S-derived promoter gave very low enzyme activity levels (average 0.16 pkat/mg). Inclusion of an N-terminal ER-specific signal peptide from a lectin gene from Phaseolus vulgaris resulted in an average activity of 1.08 pkat/mg in the transgenic tobacco lines. The additional inclusion of a C-terminal HDEL tetrapeptide, responsible for the retention of proteins in the endoplasmic reticulum of eukaryotic cells, increased the activity to 18.6 pkat/mg. When the promotor of this construct was changed from the 35S derivative to the recently described very strong plant promoter (ocs)3mas, the activity increased further to 128.6 pkat/mg. The most active tobacco line showed activities of the introduced enzyme which exceeded those of wild-type E. coli (the source of ubiA) by a factor of 1100. These results demonstrate the efficacy of plant ER-specific signal peptides for the active expression of a prokaryotic membrane protein in plants.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1023/A:1026507803067
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