Expression of recombinant growth hormone in Escherichia coli: effect of the region between the Shine-Dalgarno sequence and the ATG initiation codon.
We constructed a synthetic Escherichia coli expression system in which various promoter elements can be changed easily. In this study we investigated the effect of a number of portable Shine-Dalgarno regions (SD regions) on the synthesis of two modified recombinant human growth hormones (hGH). The production of these modified hGH was measured during exponential growth and after the bacteria had reached stationary phase. The results show that the optimal distance between the SD region (AGGAGG) and the ATG start codon is approximately 11 nucleotides. However, the nucleotide sequence in this region also influences expression: 6-10 adenines result in comparable expression levels despite the varying lengths. Two overlapping SD regions reduce expression of the growth hormones considerably, whereas two potential ATG start codons do not affect expression. Having a SD-ATG region partly or totally complementary to the 5' end of the 16S ribosomal RNA does not alter translation efficiency. Estimation of the delta G values for the association between the 16S rRNA and the ribosome-binding region suggests that these are not indicators of expression efficiency.
Dalbøge
H
H
Nordisk Gentofte, Denmark.
Carlsen
S
S
Jensen
E B
EB
Christensen
T
T
Dahl
H H
HH
D001483:Base Sequence; D003062:Codon; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D005796:Genes; D013006:Growth Hormone / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis*
D003062:Codon; D011994:Recombinant Proteins; D013006:Growth Hormone