Congratulations to Vijayendra Bhargava Nemmaru, Jenna Douglass, Antonio De Chellis, Srivatsan Shankar, Alina Thokkadam, Allan Wang, as well as our long-time collaborator at NREL (John Yarbrough), on publishing a research paper in Frontiers in Energy Research! Check out link for accessing the just accepted paper online. This was as invited article for a special journal research topic issue titled “10 Years of Frontiers in Energy Research”.
Briefly, in this study, supercharged cellulases were computationally designed using Rosetta, recombinantly expressed in E. coli, purified, and then systematically tested against various pretreated lignocellulosic biomass substrates. Although the original wild-type cellulase enzyme showed greater activity compared to both negatively and positively supercharged enzymes towards pretreated biomass without any thermal deactivation, thermal denaturation of proteins upon exposure to high temperatures prior to enzymatic assays identified two negatively supercharged constructs that outperformed the wild-type enzyme, showing ~3 to 4-fold higher activity than the control wild-type. To better understand the causal factor of reduced supercharged enzyme activity for some constructs, we also performed hydrolysis assays on purified substrates in the presence/absence of isolated lignin as well as conducted protein thermal stability assays. Overall, our work showed how a combination of impaired cellulose binding and lower thermal stability was the root cause of reduced hydrolytic activity of some cellulolytic enzyme designs. This work is a nice complement to another cellulase supercharging paper that we recently published on this topic in another journal this year as well.
Great work team on systematically demonstrating the rules of cellulolytic enzyme design using various protein ‘supercharging’ approaches for more efficient lignocellulose deconstruction for cellulosic biofuels/bioproducts manufacturing!
Thanks for funding/support over the years from the National Science Foundation (NSF) U.S. Department of Energy (DOE), Rutgers School of Engineering, ORAU, and the National Renewable Energy Laboratory. Also, special thanks to Rebecca Ong and Bruce E. Dale labs for provision of pretreated biomass samples over the course of the project.