SYSTEM FOR EXPRESSING AND TRANSPORTING RECOMBINANT PROTEINS TO THE ESCHERICHIA COLI PERIPLASM USING THE TAT (TWIN-ARGININE TRANSLOCATION) SECRETION PATHWAY.

摘要

The present invention describes an expression vector which contains a signal peptide using the Tat secretion pathway as an alternative of the Sec secretion system for the transport of cytoquinine and further proteins of biotechnological interest to Escherichia coli periplasm. The novel system consists in using the signal peptide of penicillin acylase (SPpac) which is mutated and fussed to the synthetic human interferon-y gene (hINF-y) for the transport of the protein to the Escherichia coli periplasm using the Tat pathway. The mutated SPpac includes the NdeI site at the 5¿ end, which is contained in the expression vector pEMR, at the 3¿ end the tryptophan codon being exchanged by the serine codon, thus generating the restriction site Hind III. The Leucine and Alanine codons are modified so as to generate the NheI site. Thus, two sites for restricting the cloning and fusion within a gene phase of homologous or heterologous proteins are generated. Nowadays there is no commercial vector available for the expression and transport of proteins, which include signal peptides using the Tat pathway.