CPC C07K 14/34 (2013.01) [C07K 1/22 (2013.01); C12N 15/70 (2013.01); C12N 15/902 (2013.01); C07K 2319/034 (2013.01); C07K 2319/55 (2013.01); C12N 2500/24 (2013.01); C12N 2500/34 (2013.01); C12N 2500/35 (2013.01); C12N 2500/74 (2013.01)] | 17 Claims |
1. An expression system for producing a Diphtheria toxin polypeptide or a mutated form thereof, the expression system comprising:
an Escherichia coli (E. coli) cell comprising a heterologous nucleic acid construct comprising:
(i) a rhamnose inducible promoter sequence; and
(ii) an expression sequence, said expression sequence comprising a first portion and a second portion, said first portion comprising a nucleotide sequence that encodes a periplasmic secretion signal linked to the 5′ end of the second portion, and said second portion comprising a nucleotide sequence that encodes said Diphtheria toxin polypeptide or mutated form thereof,
wherein said expression sequence is operatively linked to said rhamnose inducible promoter sequence;
wherein said E. coli cell is unable to use rhamnose as a carbon source; and
wherein:
said E. coli cell expresses a leucine/isoleucine/valine transporter subunit (LivK) fused to an affinity tag;
said E. coli cell comprises a defective livK gene;
said E. coli cell expresses a maltose transporter subunit (MalE) fused to an affinity tag;
said E. coli cell comprises a defective malE gene;
said E. coli cell expresses a LivK fused to an affinity tag and a MalE fused to an affinity tag;
said E. coli cell comprises a defective livK gene and a defective malE gene said E. coli cell expresses a LivK fused to an affinity tag and comprises a defective malE gene; or
said E. coli cell comprises a defective livK gene and expresses a MalE fused to an affinity tag.
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