Rare codon priority and its position specificity at the 5' of the gene modulates heterologous protein expression in Escherichia coli.
Rare codons and their effects in heterologous protein expression in Escherichia coli were addressed by many investigators. Here, we propose that not all rare codons of a foreign gene have negative effect but selective codon among them and its specific position in the downstream of the start codon modulates the expression. In our study, streptokinase (47 kDa), encoded by skc gene of Streptococcus equisimilis was expressed in E.coli. The analysis of relative codon frequency of skc gene in E.coli reveals the presence of 30% of rare codons in it. Nevertheless, E.coli managed to yield over-expression of this target protein. To explore the codon bias in expression, we have introduced the selective AGG codon at different positions of skc gene such as +2,+3,+5,+8,+9 and +11. The results revealed that at +2 position "AGG" aided over-expression while shifting to +3 and +5 positions it rendered nil expression. In contrary, shifting of AGG codon to later positions like +9 and +11 the inhibitory effect was reversed and resulted in over-expression. The effect of 'AGG' rare codon was further studied in GFP expression. In conclusion, besides the choice of rare codons, their precise positions in the foreign gene dictate the level of protein expression.
Thangadurai
Chinnathambi
C
Genetic Engineering Unit, Centre for Biotechnology, Sadar Patel Road, Anna University, Guindy, Chennai-600 025, India.
Suthakaran
Pichaimuthu
P
Barfal
Pankaj
P
Anandaraj
Balaiah
B
Pradhan
Satya Narayan
SN
Ramalingam
Subramanian
S
Murugan
Vadivel
V
D024506:5' Flanking Region; D001483:Base Sequence; D003062:Codon / Q000235:genetics*; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005818:Genetic Engineering; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D014176:Protein Biosynthesis / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D013291:Streptococcus / Q000201:enzymology / Q000235:genetics; D013300:Streptokinase / Q000235:genetics
D003062:Codon; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D013300:Streptokinase