title: Optimization of translation profiles enhances protein expression and solubility.
abstract: mRNA is translated with a non-uniform speed that actively coordinates co-translational folding of protein domains. Using structure-based homology we identified the structural domains in epoxide hydrolases (EHs) and introduced slow-translating codons to delineate the translation of single domains. These changes in translation speed dramatically improved the solubility of two EHs of metagenomic origin in Escherichia coli. Conversely, the importance of transient attenuation for the folding, and consequently solubility, of EH was evidenced with a member of the EH family from Agrobacterium radiobacter, which partitions in the soluble fraction when expressed in E. coli. Synonymous substitutions of codons shaping the slow-transiting regions to fast-translating codons render this protein insoluble. Furthermore, we show that low protein yield can be enhanced by decreasing the free folding energy of the initial 5'-coding region, which can disrupt mRNA secondary structure and enhance ribosomal loading. This study provides direct experimental evidence that mRNA is not a mere messenger for translation of codons into amino acids but bears an additional layer of information for folding, solubility and expression level of the encoded protein. Furthermore, it provides a general frame on how to modulate and fine-tune gene expression of a target protein.
authors.affiliation: Institute of Biochemistry and Biology, University of Potsdam, Potsdam, Germany.
authors.forename: Anne-Katrin
authors.initials: AK
authors.lastname: Hess
authors.affiliation: Institute of Biochemistry and Biology, University of Potsdam, Potsdam, Germany.
authors.forename: Paul
authors.initials: P
authors.lastname: Saffert
authors.affiliation: BRAIN, Zwingenberg, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Liebeton
authors.affiliation: Institute of Biochemistry and Biology, University of Potsdam, Potsdam, Germany; Biochemistry, Department of Chemistry and Biochemistry, University of Hamburg, Hamburg, Germany.
authors.forename: Zoya
authors.initials: Z
authors.lastname: Ignatova
mesh_terms: D016960:Agrobacterium tumefaciens / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004851:Epoxide Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D054892:Metagenome; D008958:Models, Molecular; D014176:Protein Biosynthesis; D017510:Protein Folding; D059370:RNA Folding; D012329:RNA, Bacterial / Q000737:chemistry; D012333:RNA, Messenger / Q000737:chemistry; D000069456:Silent Mutation; D012995:Solubility; D040681:Structural Homology, Protein
chemical_list: D001426:Bacterial Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D004851:Epoxide Hydrolases