Multiple Copies of the Fusion Gene cflyC-mzfDB3 Enhance the Expression of a Hybrid Antimicrobial Peptide in Pichia pastoris

摘要: The codon-optimized zebrafish β-defensin 3 mature peptide gene mzfDB3 and channel catfish c-type lysozyme cflyC were used to design the fusion cflyC-mzfDB3. In protein cflyC-mzfDB3, 4×Gly flexible amino acid linker with an enterokinase cleavage site DDDDK was designed link C-terminus of cflyC, which potentially contributes digestion recombinant cflyC-mzfDB3 by endogenous enterokinases. expressed in Pichia pastoris X-33 using expression vectors harboring 1-, 2-, or 4-copies, respectively, cassette. copy numbers P. transformants quantified real-time quantitative PCR. Their yields showed that level 4-copy transformant 2.67-fold higher than 1-copy transformant, demonstrating increase number resulted increased expression. culture medium supernatant containing exhibited antibacterial activity against Gram-positive Listeria monocytogenes Gram-negative Pseudomonas aeruginosa, indicating there may be a synergistic effect on peptide’s activity. addition, cell-free potential candidate for further development as natural hybrid antimicrobial solution.