Super microbes
for biofuel production

Considering the high amounts of CO2 produced globally from burning fossil fuels, producing fuel from CO2 would be like turning lead into gold. Bacteria and yeast can carry out this process, but the more ethanol they produce, the more their ethanol-producing capabilities are limited. Researchers at Tianjin University and the Shanghai Institutes for Biological Sciences of the Chinese Academy of Sciences in China recently reported in the journal Molecular & Cellular Proteomics a new target for genetic engineering that could increase microbial tolerance to ethanol.

Most bioethanol is produced by bacteria and yeast that derive their energy from sugars obtained from crops like corn, sparking the concern that crops are going toward biofuel production instead of food production. Cyanobacteria are attractive alternatives to the sugar-using bacteria because they obtain their energy through photosynthesis and use CO2 to produce ethanol. However, cyanobacteria are very sensitive to ethanol: A marginal amount of ethanol can slow their growth dramatically.

Ethanol, in general, is toxic to cells, but genetic engineering of sugar-using bacteria and yeast has created strains that have higher tolerance to ethanol. Several studies have suggested that these microbes employ several lines of defense to handle the excess biofuel products they produce.  Manipulating the genes that regulate transcription could control this response and confer tolerance more readily than targeting individual metabolic genes. Targeting genes that control transcription has increased significantly ethanol tolerance and the efficiency of ethanol production in sugar-using microorganisms. The research collaboration led by Weiwen Zhang sought to determine if this strategy could be used for cyanobacteria.

Using the model cyanobacterium strain, the group previously had identified by proteomics and transcriptomic analyses three proteins involved in regulating transcription, Sll0792, Sll0794 and Sll1423, whose expression is influenced by ethanol exposure. In this new study, the investigators created mutant strains that were missing those genes and grew the mutant bacteria under normal conditions or in 1.5 percent ethanol. The sll0794- and sll1423-deficient cells did not grow more slowly in ethanol, but the sll0794-deficient cells did, suggesting that disrupting the sll0794 gene altered the abundance of proteins that were important for tolerating ethanol.

The researchers then used proteomics analysis to identify which proteins were expressed differently in sll0794-deficient cells when those cells were exposed to ethanol. The investigators selected the genes that changed the most and determined if sll0749 directly controlled the transcription of those genes. They found that sll0794 bound to the promoter regions of three of the genes: sll1514, which corresponds to a 16.6 kDa small heat shock protein; slr1838, which corresponds to a carbon dioxide concentrating mechanism protein CcmK; and slr1512, which corresponds to a putative sodium-dependent bicarbonate transporter.

This is the first study to identify a specific transcriptional regulatory gene, sll0794, as a target for genetically engineering cyanobacterium for ethanol tolerance. Manipulating this gene could result in strains that can better withstand and, as a result, more efficiently produce ethanol. By overcoming the ethanol sensitivity, scientists will be closer to realizing the use of cyanobacteria to recycle CO2 to form new fuel.

Maggie Kuo Maggie Kuo is an intern at ASBMB Today and a Ph.D. candidate in biomedical engineering at Johns Hopkins University.