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Directed evolution methods for improving polypeptide folding, solubility and stability

United States Patent Application

20090142820
A1
View the Complete Application at the US Patent & Trademark Office
Los Alamos National Laboratory - Visit the Technology Transfer Division Website
The invention provides directed evolution methods for improving the folding, solubility and stability (including thermostability) characteristics of polypeptides. In one aspect, the invention provides a method for generating folding and stability-enhanced variants of proteins, including but not limited to fluorescent proteins, chromophoric proteins and enzymes. In another aspect, the invention provides methods for generating thermostable variants of a target protein or polypeptide via an internal destabilization baiting strategy. Internally destabilization a protein of interest is achieved by inserting a heterologous, folding-destabilizing sequence (folding interference domain) within DNA encoding the protein of interest, evolving the protein sequences adjacent to the heterologous insertion to overcome the destabilization (using any number of mutagenesis methods), thereby creating a library of variants. The variants in the library are expressed, and those with enhanced folding characteristics selected.
Bradbury, Andrew M. (Santa Fe, NM), Kiss, Csaba (Los Alamos, NM), Waldo, Geoffrey S. (Santa Fe, NM)
Los Alamos National Security
12/ 286,967
October 2, 2008
STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT [0002] This invention was made with government support under Contract No. DE-AC52-06 NA 25396 awarded by the U.S. Department of Energy. The government has certain rights in the invention.