Combinatorial discovery of enzymes with utility in biomass transformation
AssigneeWisconsin Alumni Research Foundation
InventorsBrian G. Fox, Nathaniel L. Elsen
Life of this patent
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Abstract
Methods for the cell-free identification of polypeptide and polypeptide combinations with utility in biomass transformation, as well as specific novel polypeptides and cell-free systems containing polypeptide combinations discovered by such methods are disclosed.
Claims26
A method for identifying a polypeptide that modulates biomass transformation comprising:
- (a) transcribing a nucleic acid fragment to make the corresponding mRNA;
- (b) translating the mRNA in a cell-free environment comprising a cell-free extract to produce a polypeptide;
- (c) contacting the polypeptide with biomass;
- and
- (d) assaying the effect of the polypeptide on transformation of the biomass to determine whether the polypeptide modulates biomass transformation.
A polypeptide comprising one or more of the amino acid sequences of the fusion proteins CelAcc_CBM (SEQ ID NO:1), CelKcc_CBM (SEQ ID NO:2), CelLcc_CBM (SEQ ID NO:3), CelRcc_CBM (SEQ ID NO:4), ChiAcc_CBM (SEQ ID NO:5), LicBcc_CBM (SEQ ID NO:6), ManAcc_CBM (SEQ ID NO:7), XynCcc_CBM (SEQ ID NO:8), or XynYcc_CBM (SEQ ID NO:9).
A nucleic acid comprising a sequence coding for one or more of the amino acid sequences set forth in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, or SEQ ID NO:9.
A cell-free expression system comprising:
- (a) a cell-free extract for synthesizing a desired target polypeptide;
- (b) a nucleotide sequence encoding one or more of the amino acid sequences set forth in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, or SEQ ID NO:9.
A cell-free expression system comprising:
- (a) a cell-free extract for synthesizing a desired target polypeptide;
- (b) a first nucleotide sequence encoding a first fusion protein, the first fusion protein comprising a first cohesin domain and a first biomass binding domain;
- and
- (c) a second nucleotide sequence encoding a second fusion protein, the second fusion protein comprising a first dockerin domain and a first target polypeptide that has a biological activity of catalyzing biomass transformation;
- wherein, when the first and second fusion proteins are expressed, the first cohesin domain can associate with the first dockerin domain.
Dependencies are inferred from the claim text.