Paul Hudson.

Graduate Student
University of California, Berkeley

B.S. Chemical Engineering, 2004
North Carolina State University

Co-Advisor: Jeffrey Reimer, Ph.D.


Office Location: 497A Tan Hall
Office Telephone: 510-643-8340
Office Fax: 510-643-1228

Activation mechanisms for enzymes in non-aqueous media

The hallmark of enzymatic catalysis is the unrivaled chemo-, regio- and stereoselectivity displayed toward complex substrates. The removal of enzymes from their natural, aqueous environs enables the extension of these selectivities toward new substrates and the use of enzymes in non-aqueous media is becoming an attractive tool in chemical synthesis. However, while the promise of fresh and exciting uses for enzymes is alluring, reduced reaction rates in non-aqueous media looms as a large barrier to the widespread implementation of non-aqueous biocatalysis on the industrial scale.

Traditional explanations of the deleterious effects of organic solvents include the rigidifcation and reduced conformational mobility in low dielectric media, the partial loss of tertiary and secondary structure, and the subsequent disruption of the active-site electronic environment (for a recent review, see [1]). The methods used to increase enzyme activity in organic solvents range from simple addition of water [2] or co-lyophilization with inert salts [3] to protein engineering techniques such as directed evolution and rational mutagenesis.

In the Clark and Reimer labs, I utilize the methods of biophysical spectroscopy with contemporary enzyme engineering so as to craft proteins to be more catalytically active in non-native environments. My work to date falls into two categories: examination with 19F NMR of several properties of the enzyme subtilisin’s active site when the enzyme was dissolved in organic solvents and at various hydration levels. These active-site specific studies revealed that active-site polarity and motion on a particular timescale are important for catalysis and that hydration of the enzyme promotes both [2]. The other part of my project utilizes 2D NMR methods to interrogate the backbone of a homologous protein to determine which amino acids are most affected by the presence of organic solvents. These spectra guide us in our engineering approach by suggesting which amino acid mutations may improve catalytic activity in organic solvents.

[1] Hudson EP, Eppler RK, Clark DS. “Biocatalysis in semi-aqueous and nearly anhydrous conditions.” Curr. Opin. Biotech. 2005, 16: 637-643

[2] Hudson EP, Eppler RK, Beaudoin JM, Dordick JS, Reimer JA, Clark DS. “Active-site motions and polarity enhance catalytic turnover of hydrated subtilisin dissolved in organic solvents.” Journal of the American Chemical Society 2009 In press. DOI 10.1021/ja806996q

[3] Eppler RK, Hudson EP, Chase SD, Dordick JS, Reimer JA, Clark DS. “Biocatalyst activity in non-aqueous environments correlates with centisecond-range protein motions.” Proceedings of the National Academy of Sciences USA. 2008 105: 15672-15677