Optimized bglA gene from Cellulomonas flavigena expressed in Saccharomyces cerevisiae for ethanol production in cellobiose-rich sugar cane bagasse saccharified
Tuesday, April 29, 2014
Exhibit/Poster Hall, lower level (Hilton Clearwater Beach)
Francisco Javier Ríos-Fránquez1, Ana C. Ramos-Valdivia1, Héctor M. Poggi-Varaldo1, Jaime García-Mena2, Alfredo Martínez-Jiménez3 and Ma. Teresa Ponce-Noyola1, (1)Biotechnology and Bioengineering, CINVESTAV, Mexico, D.F., Mexico, (2)Genetics and Molecular Biology, CINVESTAV, Mexico, D.F., Mexico, (3)Cell Engineering and Biocatalysis, IBT - UNAM, Cuernavaca, Mexico
Bioethanol is one of the main alternative biofuels currently used in vehicles. However in Mexico should be obtained from the fermentation of saccharified agricultural residues. But this technology actually is expensive and needs to be optimized to make it profitable.

We have studied the use of holocellulases from Cellulomonas flavigena to saccharify agricultural waste. This bacterium expresses all its cellulases extracellularly, except its β-glucosidase, which remains in the periplasmic space, requiring the lysis of the bacteria to release it out of the cells. Therefore the saccharified obtained with extracellular enzymatic extracts from C. flavigena are rich in cellobiose.

An alternative to produce bioethanol from saccharified obtained with cellulases from C. flavigena, is to express recombinant β-glucosidase in Saccharomyces cerevisiae to grow and ferment cellobiose-rich saccharified.

We identified and sequenced bglA gene from C. flavigena encoding β-glucosidase, and expressed in S. cerevisiae using the plasmid pYEX-S1. However the levels of expression of β-glucosidase and production of ethanol in cellobiose with this recombinant yeast were very low.

To enhance expression of C. flavigena β-glucosidase, we optimized the bglA gene by adapting the codon usage to the preferred use for S. cerevisiae. The optimized gene was cloned into the expression vector pYEX-S1 and transformed into S. cerevisiae competent cells. The optimized gene improves the extracellular β-glucosidase activity and also the yields of ethanol from recombinant yeast growing in cellobiose and saccharified sugar cane bagasse.