Harvard College

The Broad Institute, Inc.

Tina Wang

David R Liu

Ahmed Hussein Badran

Some aspects of this disclosure relate to strategies, systems, methods, compositions, and kits that are useful for production (e.g., evolution) of cytidine deaminase protein variants that are characterized by increased soluble expression and/or stability relative to the wild-type cytidine deaminase protein from which they are evolved. In some embodiments, evolved cytidine deaminase variants described by the disclosure are useful for incorporation into targeted nucleic acid editing proteins, for example in fusion proteins with a Cas9 domain or variant thereof.

Evolution of cytidine deaminases

Some aspects of this disclosure relate to systems, apparatuses, compositions (e.g., isolated nucleic acids and vectors), and methods for improving the stability and/or solubility of proteins evolved using phage-assisted continuous evolution (PACE). In some embodiments, vectors described herein comprise nucleic acids encoding selection systems (e.g., positive and/or negative selection systems) that link expression of genes required for production of infectious phage particles to a desirable physiochemical (e.g., stability or solubility) and/or desired function of an evolved protein.

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Cambridge, MA, United States of America

Tina Wang