Methods and compositions for polypeptide engineering
AssigneeMaxygen, Inc.
InventorsPhillip A. Patten, Willem P. C. Stemmer
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Abstract
Methods are provided for the evolution of proteins of industrial and pharmaceutical interest, including methods for effecting recombination and selection. Compositions produced by these methods are also disclosed.
Claims37
A method for evolving a polypeptide to comprise an improved serum half-life or stability, the polypeptide being encoded by a DNA substrate molecule, the method comprising:
- (a) providing a library of variants of the DNA substrate molecule;
- (b) transforming the variants of (a) into host cells;
- (c) expressing the variants of (b) to provide polypeptide expression products;
- (d) screening or selecting from the polypeptide expression products of (c) an evolved polypeptide comprising the improved serum half-life or stability relative to the polypeptide encoded by the DNA substrate molecule in vivo;
- (e) recovering an evolved DNA substrate molecule that encodes the evolved polypeptide of (d);
- and,
- (f) expressing the evolved DNA substrate molecule, thereby providing the evolved polypeptide comprising the improved serum half-life or stability.
A method for evolving a DNA substrate molecule encoding a polypeptide comprising an improved serum half-life or stability, the method comprising:
- (a) transfecting a host cell with at least first and second DNA substrate molecules wherein the first and second DNA substrate molecules are recombined in the host cell;
- (b) expressing the recombined DNA substrate molecules of (a) to provide polypeptide expression products;
- (c) screening or selecting from the polypeptide expression products of (b) an evolved polypeptide comprising the improved serum half-life or stability relative to the polypeptide encoded by the DNA substrate molecule;
- and,
- (d) derivatizing the evolved polypeptide of (c) with a half-life extending moiety.
A method for evolving a DNA substrate molecule encoding a polypeptide comprising an improved serum half-life or stability, the method comprising:
- (a) transfecting a host cell with at least first and second DNA substrate molecules wherein the at least first and second DNA substrate molecules are recombined in the host cell;
- (b) expressing the recombined DNA substrate molecules of (a) to provide polypeptide expression products;
- (c) derivatizing the polypeptide expression products of (b) with a half-life extending moiety to provide derivatized polypeptide expression products;
- and,
- (d) screening or selecting from the derivatized polypeptide expression products of an evolved polypeptide comprising the improved serum half-life or stability relative to the polypeptide encoded by the DNA substrate molecule derivatized with the half-life extending moiety.
A method for evolving polypeptides to comprise an improved serum half-life or stability, the polypeptides being encoded by DNA substrate molecules, the method comprising:
- (a) providing a library of variants of the DNA substrate molecules;
- (b) transforming the variants of (a) into host cells;
- (c) expressing the variants of (b) to provide polypeptide expression products;
- (d) screening or selecting from the polypeptide expression products of (c) evolved polypeptides comprising the improved serum half-life or stability relative to the polypeptides encoded by the DNA substrate molecules;
- and,
- (e) derivatizing the evolved polypeptides of (d) with a half-life extending moiety in order to further extend the in half-life or stability of the evolved polypeptides.
A method for evolving polypeptides to comprise an improved serum half-life or stability, the polypeptides being encoded by DNA substrate molecules, the method comprising:
- (a) providing a library of variants of the DNA substrate molecules;
- (b) transforming the variants of (a) into host cells;
- (c) expressing the variants of (b) to provide polypeptide expression products;
- (d) derivatizing the polypeptide expression products of (c) with a half-life extending moiety to provide derivatized polypeptide expression products;
- and,
- (e) screening or selecting from the derivatized polypeptide expression products of (d) evolved polypeptides comprising the improve serum half-life or stability relative to the polypeptides encoded by the DNA substrate molecules derivatized with the half-life extending moiety.
Dependencies are inferred from the claim text.