Reengineering mRNA Primary Structure for Enhanced Protein Production
AssigneeThe Scripps Research Institute
InventorsVincent P. Mauro, Stephen A. Chappell, Wei Zhou, Gerald M. Edelman
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Abstract
Described herein are rules to modify natural mRNAs or to engineer synthetic mRNAs to increase their translation efficiencies. These rules describe modifications to mRNA coding and 3′ UTR sequences intended to enhance protein synthesis by: 1) decreasing ribosomal diversion via AUG or non-canonical initiation codons in coding sequences, and/or 2) by evading miRNA-mediated down-regulation by eliminating one or more miRNA binding sites in coding sequences.
Claims23
A method of improving full-length protein expression efficiency comprising:
- a) providing a polynucleotide comprising:
- i) a coding sequence for the protein;
- ii) a primary initiation codon that is upstream of the coding sequence;
- and
- iii) one or more secondary initiation codons located within the coding sequence;
- and
- b) mutating one or more secondary initiation codons, wherein the mutation results in a decrease in initiation of protein synthesis at the one or more secondary initiation codons resulting in a reduction of ribosomal diversion away from the primary initiation codon,
- thereby increasing full-length protein expression efficiency.
A method of improving full-length protein expression efficiency comprising:
- a) providing a polynucleotide sequence comprising a coding sequence for the protein and one or more miRNA binding sites located within the coding sequence;
- and
- b) mutating the one or more miRNA binding sites, wherein the mutation results in a decrease in miRNA binding at the one or more miRNA binding sites resulting in a reduction of miRNA-mediated down regulation of protein translation, thereby increasing full-length protein expression efficiency.
Dependencies are inferred from the claim text.