Sieber, Volker and Martinez, Carlos A. and Arnold, Frances H. (2001) Libraries of hybrid proteins from distantly related sequences. Nature Biotechnology, 19 (5). pp. 456-460. ISSN 1087-0156. doi:10.1038/88129. https://resolver.caltech.edu/CaltechAUTHORS:20181113-135322855
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Abstract
We introduce a method for sequence homology–independent protein recombination (SHIPREC) that can create libraries of single-crossover hybrids of unrelated or distantly related proteins. The method maintains the proper sequence alignment between the parents and introduces crossovers mainly at structurally related sites distributed over the aligned sequences. We used SHIPREC to create a library of interspecies hybrids of a membrane-associated human cytochrome P450 (1A2) and the heme domain of a soluble bacterial P450 (BM3). By fusing the hybrid gene library to the gene for chloramphenicol acetyl transferase (CAT), we were able to select for soluble and properly folded protein variants. Screening for 1A2 activity (deethylation of 7-ethoxyresorufin) identified two functional P450 hybrids that were more soluble in the bacterial cytoplasm than the wild-type 1A2 enzyme.
| Item Type: | Article | |||||||||
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| Additional Information: | © 2001 Nature Publishing Group. Received 21 November 2000; Accepted 31 January 2001; Published 01 May 2001. V.S. was supported by a research fellowship from the Deutsche Forschungsgemeinschaft (DGF). This work was funded by Maxygen Inc. | |||||||||
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| Issue or Number: | 5 | |||||||||
| DOI: | 10.1038/88129 | |||||||||
| Record Number: | CaltechAUTHORS:20181113-135322855 | |||||||||
| Persistent URL: | https://resolver.caltech.edu/CaltechAUTHORS:20181113-135322855 | |||||||||
| Usage Policy: | No commercial reproduction, distribution, display or performance rights in this work are provided. | |||||||||
| ID Code: | 90875 | |||||||||
| Collection: | CaltechAUTHORS | |||||||||
| Deposited By: | Tony Diaz | |||||||||
| Deposited On: | 13 Nov 2018 22:41 | |||||||||
| Last Modified: | 16 Nov 2021 03:36 |
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