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Applied and Environmental Microbiology, May 2008, p. 3182-3188, Vol. 74, No. 10
0099-2240/08/$08.00+0     doi:10.1128/AEM.02858-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Engineering and Analysis of a Saccharomyces cerevisiae Strain That Uses Formaldehyde as an Auxiliary Substrate

Molecular Cell Biology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, P.O. Box 14, 9750 AA Haren, The Netherlands,¹ Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands,² Kluyver Centre for Genomics of Industrial Fermentation, Julianalaan 67, 2628 BC Delft, The Netherlands³

Received 19 December 2007/ Accepted 20 March 2008

We demonstrated that formaldehyde can be efficiently coutilized by an engineered Saccharomyces cerevisiae strain that expresses Hansenula polymorpha genes encoding formaldehyde dehydrogenase (FLD1) and formate dehydrogenase (FMD), in contrast to wild-type strains. Initial chemostat experiments showed that the engineered strain coutilized formaldehyde with glucose, but these mixed-substrate cultures failed to reach steady-state conditions and did not exhibit an increased biomass yield on glucose. Subsequent transcriptome analyses of chemostat cultures of the engineered strain, grown on glucose-formaldehyde mixtures, indicated that the presence of formaldehyde in the feed caused biotin limitations. Further transcriptome analysis demonstrated that this biotin inactivation was prevented by using separate formaldehyde and vitamin feeds. Using this approach, steady-state glucose-limited chemostat cultures were obtained that coutilized glucose and formaldehyde. Coutilization of formaldehyde under these conditions resulted in an enhanced biomass yield of the glucose-limited cultures. The biomass yield was quantitatively consistent with the use of formaldehyde as an auxiliary substrate that generates NADH and subsequently, via oxidative phosphorylation, ATP. On an electron pair basis, the biomass yield increase observed with formaldehyde was larger than that observed previously for formate, which is tentatively explained by different modes of formate and formaldehyde transport in S. cerevisiae.

Published ahead of print on 31 March 2008.