title: Codon optimization increases human kallistatin expression in Escherichia coli.
abstract: A unique serpin, kallistatin, displays vasodilatory, antiangiogenic, anti-inflammatory, and antioxidant activity. Difficulty and low efficacy of obtaining recombinant kallistatin limit the wide investigation of its biological and pathological function. The present study employed a codon optimization algorithm to redesign the kallistatin gene and achieved a high yield of recombinant kallistatin protein. The kallistatin codons were redesigned for a more suitable Escherichia coli host without altering amino acids. Base composition and GC% content were compared between synthetic optimized kallistatin (opti-kallistatin) and wild-type kallistatin (wt-kallistatin). Both opti-kallistatin and wt-kallistatin were purified using Ni-NTA His-binding resins through fast protein liquid chromatography (FPLC). The identity and purity of kallistatin were confirmed by Coomassie blue staining, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and Western blot analysis. The output of opti-kallistatin protein was ~2-fold increase (2.09 ± 0.23 mg/L) compared to wt-kallistatin (1.05 ± 0.2 mg/L). These results suggest that more common codon optimization in the E. coli host significantly increases the yield of heterologous human protein yields. This approach will remarkably facilitate the further investigation of kallistatin in vitro and in vivo.
authors.affiliation: Department of Biochemistry, Zhongshan Medical School, Sun Yat-sen University, Guangzhou, Guangdong Province, China.
authors.forename: Zhiyu
authors.initials: Z
authors.lastname: Dai
authors.forename: Yifei
authors.initials: Y
authors.lastname: Chen
authors.forename: Weiwei
authors.initials: W
authors.lastname: Qi
authors.forename: Lijun
authors.initials: L
authors.lastname: Huang
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhang
authors.forename: Ti
authors.initials: T
authors.lastname: Zhou
authors.forename: Xia
authors.initials: X
authors.lastname: Yang
authors.forename: Guoquan
authors.initials: G
authors.lastname: Gao
mesh_terms: D001483:Base Sequence; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003062:Codon; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D015843:Serpins / Q000096:biosynthesis / Q000235:genetics* / Q000494:pharmacology
chemical_list: D003062:Codon; D003470:Culture Media; D011994:Recombinant Proteins; D015843:Serpins; C051324:kallistatin