Optimization of DsRed production in Escherichia coli: effect of ribosome binding site sequestration on translation efficiency.
DsRed-Express, a popular reporter protein, cannot be expressed in Escherichia coli using a consensus ribosome binding site (RBS) potentially due to basepairing in the RBS that inhibits translation initiation. Saturation mutagenesis was used to probe for a gene sequence that minimized basepairing in the RBS while maintaining the same spectral properties and maturation characteristics as DsRed-Express. The new DsRed, designated here as RFP(EC) for E. coli optimized red fluorescent protein, fluoresces 2.5 times greater than DsRed-Express when expressed from the same vector.
Pfleger
Brian F
BF
Department of Chemical Engineering, University of California Berkeley, California, USA.
Fawzi
Nicolas J
NJ
Keasling
Jay D
JD
D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D017930:Genes, Reporter / Q000235:genetics; D024861:Genetic Enhancement / Q000379:methods*; D008164:Luminescent Proteins / Q000096:biosynthesis* / Q000235:genetics; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods*; D046188:Protein Modification, Translational / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis; D012270:Ribosomes / Q000378:metabolism*
D008164:Luminescent Proteins; D011994:Recombinant Proteins; C413663:fluorescent protein 583