title: N-terminal SKIK peptide tag markedly improves expression of difficult-to-express proteins in Escherichia coli and Saccharomyces cerevisiae.
abstract: Despite advances in microbial protein expression systems, low production of proteins remains a great concern for some genes. Here we report that the insertion of a short peptide tag, consisting of Ser-Lys-Ile-Lys (SKIK), adjacent to the start codon of genes encoding difficult-to-express proteins can increase protein expression in Escherichia coli and Saccharomyces cerevisiae. Protein expression levels of a mouse monoclonal antibody (mAb), rabbit mAbs obtained from clonal B cells, and an artificially designed peptide were significantly increased simply by the addition of the SKIK tag in E. coli systems. In particular, a ∼30-fold increase in protein production was observed for the mouse mAb, and the artificially designed peptide band became detectable in sodium dodecyl sulfate-poly acrylamide gel electrophoresis after coomassie brilliant blue staining or western blotting on adding the SKIK tag. The tag also increased the expression of tagged proteins in S. cerevisiae and an E. coli cell-free protein synthesis system. Although the mechanism of high protein expression on addition of the tag is unclear, our findings offer great benefits to biotechnology research and industry.
authors.affiliation: Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan; Knowledge Hub Aichi, Aichi Science and Technology Foundation, Yakusa-cho, Toyota 470-0356, Japan.
authors.forename: Teruyo
authors.initials: T
authors.lastname: Ojima-Kato
authors.affiliation: Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan.
authors.forename: Satomi
authors.initials: S
authors.lastname: Nagai
authors.affiliation: Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan. Electronic address: hnakano@agr.nagoya-u.ac.jp.
authors.forename: Hideo
authors.initials: H
authors.lastname: Nakano
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D002474:Cell-Free System; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D051379:Mice; D009842:Oligopeptides / Q000235:genetics* / Q000378:metabolism; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D009842:Oligopeptides; D011994:Recombinant Proteins
keywords: Cell-free protein synthesis; Difficult-to-express protein; Escherichia coli; Increase of protein expression; N-terminal peptide tag; Saccharomyces cerevisiae