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High-level expression of the human tumor necrosis factor-alpha.

Five expression plasmids for a total chemically synthesized gene of tumor necrosis factor-alpha (TNF-alpha) were constructed, which differ in two aspects: 1) distance (D) between the SD and the initiation codon ATG; 2) energy-delta G0f298 released to form the stable secondary structure in the translation initiation region. The plasmid with a (D) of 6bp has the lowest delta G0f298 (absolute value) and showed the highest expression level, up to 60% of the total bacterial proteins. Codons usage will impose much influence on expression level, so it is reasonable and understandable that the total chemically synthesized TNF-alpha gene with the codons preferable to E. coli use gave much higher expression level than the sc-TNF-alpha in which only partial N-terminal codons were modified.