%0 Journal Article %1 Roethlisberger08NKemp %A Rothlisberger, Daniela %A Khersonsky, Olga %A Wollacott, Andrew M %A Jiang, Lin %A DeChancie, Jason %A Betker, Jamie %A Gallaher, Jasmine L %A Althoff, Eric A %A Zanghellini, Alexandre %A Dym, Orly %A Albeck, Shira %A Houk, Kendall N %A Tawfik, Dan S %A Baker, David %D 2008 %J Nature %K TobeLearning ComputDesign SimulatingTechnology %N 7192 %P 190--195 %R 10.1038/nature06879 %T Kemp elimination catalysts by computational enzyme design. %U http://dx.doi.org/10.1038/nature06879 %V 453 %X The design of new enzymes for reactions not catalysed by naturally occurring biocatalysts is a challenge for protein engineering and is a critical test of our understanding of enzyme catalysis. Here we describe the computational design of eight enzymes that use two different catalytic motifs to catalyse the Kemp elimination-a model reaction for proton transfer from carbon-with measured rate enhancements of up to 10(5) and multiple turnovers. Mutational analysis confirms that catalysis depends on the computationally designed active sites, and a high-resolution crystal structure suggests that the designs have close to atomic accuracy. Application of in vitro evolution to enhance the computational designs produced a >200-fold increase in k(cat)/K(m) (k(cat)/K(m) of 2,600 M(-1)s(-1) and k(cat)/k(uncat) of >10(6)). These results demonstrate the power of combining computational protein design with directed evolution for creating new enzymes, and we anticipate the creation of a wide range of useful new catalysts in the future.

@article{Roethlisberger08NKemp, abstract = {The design of new enzymes for reactions not catalysed by naturally occurring biocatalysts is a challenge for protein engineering and is a critical test of our understanding of enzyme catalysis. Here we describe the computational design of eight enzymes that use two different catalytic motifs to catalyse the Kemp elimination-a model reaction for proton transfer from carbon-with measured rate enhancements of up to 10(5) and multiple turnovers. Mutational analysis confirms that catalysis depends on the computationally designed active sites, and a high-resolution crystal structure suggests that the designs have close to atomic accuracy. Application of in vitro evolution to enhance the computational designs produced a >200-fold increase in k(cat)/K(m) (k(cat)/K(m) of 2,600 M(-1)s(-1) and k(cat)/k(uncat) of >10(6)). These results demonstrate the power of combining computational protein design with directed evolution for creating new enzymes, and we anticipate the creation of a wide range of useful new catalysts in the future.}, added-at = {2009-10-05T07:59:51.000+0200}, author = {Rothlisberger, Daniela and Khersonsky, Olga and Wollacott, Andrew M and Jiang, Lin and DeChancie, Jason and Betker, Jamie and Gallaher, Jasmine L and Althoff, Eric A and Zanghellini, Alexandre and Dym, Orly and Albeck, Shira and Houk, Kendall N and Tawfik, Dan S and Baker, David}, biburl = {https://www.bibsonomy.org/bibtex/264b6185c8df5a121ae1db5be57333588/zwy}, doi = {10.1038/nature06879}, file = {Roethlisberger08NKemp.pdf:Proteins/Roethlisberger08NKemp.pdf:PDF}, institution = {Department of Biochemistry, University of Washington, Seattle, Washington 98195, USA.}, interhash = {9f3d58267eeb9de51ae3b225f5030499}, intrahash = {64b6185c8df5a121ae1db5be57333588}, journal = {Nature}, keywords = {TobeLearning ComputDesign SimulatingTechnology}, language = {eng}, medline-pst = {ppublish}, month = May, number = 7192, pages = {190--195}, pdf = {Proteins/Roethlisberger08NKemp.pdf}, pii = {nature06879}, pmid = {18354394}, timestamp = {2009-10-05T07:59:51.000+0200}, title = {Kemp elimination catalysts by computational enzyme design.}, url = {http://dx.doi.org/10.1038/nature06879}, volume = 453, year = 2008 }