Abstract
BackgroundTranslational regulation played an important role in the correct folding of heterologous proteins to form bioactive conformations during biogenesis. Translational pausing coordinates protein translation and co-translational folding. Decelerating translation elongation speed has been shown to improve the soluble protein yield when expressing heterologous proteins in industrial expression hosts. However, rational redesign of translational pausing via synonymous mutations may not be feasible in many cases. Our goal was to develop a general and convenient strategy to improve heterologous protein synthesis in Pichia pastoris without mutating the expressed genes.ResultsHere, a large-scale deletion library of ribosomal protein (RP) genes was constructed for heterologous protein expression in Pichia pastoris, and 59% (16/27) RP deletants have significantly increased heterologous protein yield. This is due to the delay of 60S subunit assembly by deleting non-essential ribosomal protein genes or 60S subunit processing factors, thus globally decreased the translation elongation speed and improved the co-translational folding, without perturbing the relative transcription level and translation initiation.ConclusionGlobal decrease in the translation elongation speed by RP deletion enhanced co-translational folding efficiency of nascent chains and decreased protein aggregates to improve heterologous protein yield. A potential expression platform for efficient pharmaceutical proteins and industrial enzymes production was provided without synonymous mutation.
Highlights
Translational regulation played an important role in the correct folding of heterologous proteins to form bioactive conformations during biogenesis
Compared to 79 ribosomal protein (RP) encoded by 138 genes in S. cerevisiae [25], 77 RPs are encoded by 86 genes and the homologous proteins of Rpl27 and Rpl41 from S. cerevisiae are missing in P. pastoris (Additional file 1: Table S1)
These findings indicated that the number and composition of the RPs of two budding yeasts are totally different
Summary
Translational regulation played an important role in the correct folding of heterologous proteins to form bioactive conformations during biogenesis. Green fluorescent protein (GFP) is a eukaryotic protein containing 11 beta-sheet structures and its folding yield was significantly increased by co-translational folding in E. coli [14] Another protein, phytase (Phy) from Citrobacter amalonaticus CGMCC 1696, has 95% homology to E. coli-derived phytase which consists of one α-domain containing five α-helices and a β-hairpin, and one α/βdomain including seven β-sheets and four α-helices [15]. Phytase (Phy) from Citrobacter amalonaticus CGMCC 1696, has 95% homology to E. coli-derived phytase which consists of one α-domain containing five α-helices and a β-hairpin, and one α/βdomain including seven β-sheets and four α-helices [15] These proteins are beta-sheet rich or multi-domain and aggregation prone, which may require higher co-translational folding efficiency
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
