title: Codon Usage in Signal Sequences Affects Protein Expression and Secretion Using Baculovirus/Insect Cell Expression System.
abstract: By introducing synonymous mutations into the coding sequences of GP64sp and FibHsp signal peptides, the influences of mRNA secondary structure and codon usage of signal sequences on protein expression and secretion were investigated using baculovirus/insect cell expression system. The results showed that mRNA structural stability of the signal sequences was not correlated with the protein production and secretion levels, and FibHsp was more tolerable to codon changes than GP64sp. Codon bias analyses revealed that codons for GP64sp were well de-optimized and contained more non-optimal codons than FibHsp. Synonymous mutations in GP64sp sufficiently increased its average codon usage frequency and resulted in dramatic reduction of the activity and secretion of luciferase. Protein degradation inhibition assay with MG-132 showed that higher codon usage frequency in the signal sequence increased the production as well as the degradation of luciferase protein, indicating that the synonymous codon substitutions in the signal sequence caused misfolding of luciferase instead of slowing down the protein production. Meanwhile, we found that introduction of more non-optimal codons into FibHsp could increase the production and secretion levels of luciferase, which suggested a new strategy to improve the production of secretory proteins in insect cells.
authors.affiliation: College of Life Sciences, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.
authors.forename: Yalan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: College of Life Sciences, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.|State key laboratory of crop stress biology for arid areas, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.
authors.forename: Yuanhui
authors.initials: Y
authors.lastname: Mao
authors.affiliation: College of Life Sciences, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Xu
authors.affiliation: College of Life Sciences, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.|State key laboratory of crop stress biology for arid areas, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.
authors.forename: Shiheng
authors.initials: S
authors.lastname: Tao
authors.affiliation: College of Life Sciences, Northwest A&F University, Yangling, Shaanxi 712100, P. R. China.
authors.forename: Hongying
authors.initials: H
authors.lastname: Chen
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D020029:Base Pairing; D001483:Base Sequence; D003062:Codon / Q000235:genetics*; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000235:genetics*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D020871:RNA Stability / Q000235:genetics*; D018411:Spodoptera / Q000166:cytology / Q000235:genetics* / Q000821:virology
chemical_list: D003062:Codon; D021382:Protein Sorting Signals; D011506:Proteins; D008156:Luciferases