Enhancing activity of epsilon in Escherichia coli and Agrobacterium tumefaciens cells.
Epsilon (epsilon) sequence is a bacterial enhancer of translation found in the bacteriophage T7 gene 10. It is believed that its enhancing effect of epsilon is due to a base-pairing with the nucleotides 458-467 from the helical domain 17 of Escherichia coli 16S rRNA. To prove this we have taken advantage of the difference of this domain in Agrobacterium tumefaciens and E. coli. To evaluate the significance of nucleotide complementarity for the enhancing activity of epsilon, a series of nucleotide sequences matching either E. coli or A. tumefaciens domain 17 are cloned in a binary expression vector in front of the chloramphenicol acetyltransferase (CAT) gene. The CAT assay shows that: (i) the epsilon in combination with an SD consensus sequence increases the yield of CAT in both microorganisms over that obtained with the SD alone; (ii) the epsilon sequence complementary to the A. tumefaciens domain 17 leads to a 2.71-fold increase in the yield of CAT in homologous cells but not in E. coli cells; (iii) the yield of CAT correlates with the free energy of base-pairing with the helical domain 17 in both microorganisms.
Golshani
A
A
Department of Botany, Virology Group, University of Toronto, 25 Willcocks Street, Toronto, Ontario, M5S 3B2, Canada.
Kolev
V
V
Mironova
R
R
AbouHaidar
M G
MG
Ivanov
I G
IG
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D001483:Base Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004269:DNA, Bacterial; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli / Q000235:genetics*; D009690:Nucleic Acid Conformation; D014176:Protein Biosynthesis; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000235:genetics
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D015500:Chloramphenicol O-Acetyltransferase