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Incorporation of partial polyhedrin homology sequences (PPHS) enhances the production of cloned foreign genes in a baculovirus expression system
Author(s) -
Gong Zhaohui,
Jin Yongfeng,
Zhang Yaozhou
Publication year - 2006
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1042/ba20050163
Subject(s) - polyhedrin , gene , biology , fusion protein , homology (biology) , coding region , microbiology and biotechnology , amino acid , fusion gene , peptide sequence , expression vector , recombinant dna , sequence analysis , gene expression , genetics , spodoptera
Baculovirus expression vector systems (BEVSs) have been used extensively for high‐level expression of cloned foreign genes. In many instances, the levels of recombinant protein(s) produced in insect cells and larvae are insufficient for experimental purposes. Thus new techniques and methods are needed to increase significantly the protein expression levels in BEVS. In the present paper, we describe the incorporation of a 15 bp element derived from the 5′‐end partial sequence of the polyhedrin gene, which contains the non‐coding sequence ATAAAT and the coding sequence ATGCCGAAT, into the 5′‐end of the CTB (cholera toxin B subunit)–INS (insulin) fusion gene. With the addition of the PPHS (partial polyhedrin homology sequences), two extra amino acids (Pro‐Asn) were added to the N‐terminus of the mCTB–INS (modified CTB–INS) fusion protein. This new fusion protein was expressed in both insect cells and larvae using BEVSs. We found that the addition of PPHS enhanced 4‐fold the expression of CTB–INS in both insect cells and larvae. Further analysis revealed that the additional two amino acids in mCTB–INS did not significantly affect binding affinity for G M1 ganglioside. Therefore the PPHS can be used as a constitutive element immediately downstream of the polyhedrin promoter to induce significant increases in the expression levels of cloned foreign genes.