Abstract
BackgroundThe methylotrophic yeast Pichia pastoris is well-known for the production of a broad spectrum of functional types of heterologous proteins including enzymes, antigens, engineered antibody fragments, and next gen protein scaffolds and many transcription factors are utilized to address the burden caused by the high expression of heterologous proteins. In this article, a novel P. pastoris transcription factor currently annotated as Fhl1p, an activator of ribosome biosynthesis processing, was investigated for promoting the expression of the recombinant proteins.ResultsThe function of Fhl1p of P. pastoris for improving the expression of recombinant proteins was verified in strains expressing phytase, pectinase and mRFP, showing that the productivity was increased by 20–35%. RNA-Seq was used to study the Fhl1p regulation mechanism in detail, confirming Fhl1p involved in the regulation of rRNA processing genes, ribosomal small/large subunit biogenesis genes, Golgi vesicle transport genes, etc., which contributed to boosting the expression of foreign proteins. The overexpressed Fhl1p strain exhibited increases in the polysome and monosome levels, showing improved translation activities.ConclusionThis study illustrated that the transcription factor Fhl1p could effectively enhance recombinant protein expression in P. pastoris. Furthermore, we provided the evidence that overexpressed Fhl1p was related to more active translation state.
Highlights
The methylotrophic yeast Pichia pastoris is well-known for the production of a broad spectrum of functional types of heterologous proteins including enzymes, antigens, engineered antibody fragments, and gen protein scaffolds and many transcription factors are utilized to address the burden caused by the high expression of heterologous proteins
The PpFhl1p was compared to S. cerevisiae (Sc) Fhl1p (GenBank accession number: P39521) using DNAMAN (Lynnon Biosoft Company)
High conservation was found for the PpFhl1p residues 350–487 compared to the S. cerevisiae ScFhl1p (Fig. 1)
Summary
A novel P. pastoris transcription factor currently annotated as Fhl1p, an activator of ribosome biosynthesis processing, was investigated for promoting the expression of the recombinant proteins. Translation, a process by which a ribosome reads an mRNA template to guide protein synthesis, is critical for gene expression and costs tremendous energy [11, 12]. Improving the mRNA level of target gene is commonly applied to increase the protein expression. When optimized promoter and gene dosages are used to enhance the abundance of mRNA, target protein productivity does not linearly increase as expected [16, 17]. Few studies have been performed on regulating the translation of yeast strains when engineered to express high protein yields. It is meaningful to find a novel transcription factor to regulate translation in engineered strains for industrial production [6]
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