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Deleting the first disulphide bond in an arenicin derivative enhances its expression in Pichia pastoris.

Abstract：

:The marine antimicrobial peptide NZ17074, a variant of arenicin-3 from Arenicola marina that has broad antimicrobial activity and high bioavailability, can be designed to treat bacterial and fungal diseases. To reduce the toxicity of NZ17074, N6 was designed by replacing a cysteine in positions 3 and 20 with alanine, fused to the C-terminus of the small ubiquitin-like modifier tag (SUMO), and expressed in yeast. SUMO-N6 yielded as much as 921 mg l-1 at 72 h after induction in a fermentor and increased 1·8-fold over SUMO-NZ17074. After cleavage with 30% formic acid and purification by a Sephadex G-25 column, 9·7 mg of the recombinant peptide N6 (rN6) was obtained from one-litre fermentation broth, increasing 1·4-fold over NZ17074. Compared to NZ17074, rN6 displayed almost identical antimicrobial activity with a minimal inhibitory concentration of 0·5, 0·25-0·5, 4, 0·25-16 and 16 μg ml-1 against Escherichia, Salmonella, Pseudomonas, Staphylococcus and Streptococcus strains. Our results indicate that the first disulphide bond, Cys3-Cys20, in NZ17074 is not necessary for antimicrobial activity and that its deletion might reduce toxicity to host cells. These findings may help design new antimicrobial peptides harbouring fewer disulphide bridges and may have more potent activity. SIGNIFICANCE AND IMPACT OF THE STUDY:Disulphide bond formation is an important step in the protein expression and can also influence protein secretion. A deletion of the first disulphide bond in NZ17074 increased the secreted level of target protein, and its antimicrobial activity was almost unaffected by the deletion of the first disulphide bond. The first disulphide bond in NZ17074 is favourable for correctly forming another disulphide bond during expression but not necessary for its activity. This may help design and produce a novel class of antimicrobial peptides harbouring fewer disulphide bridges to save the cost.

journal_name

Lett Appl Microbiol

journal_title

Letters in applied microbiology

authors

Yang N,Wang X,Teng D,Mao R,Hao Y,Feng X,Wang J

doi

10.1111/lam.12770

subject

Has Abstract

pub_date

2017-09-01 00:00:00

pages

241-248

issue

eissn

0266-8254

issn

1472-765X

journal_volume

pub_type

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Deleting the first disulphide bond in an arenicin derivative enhances its expression in Pichia pastoris.