Mixed acid fermentation

Biochemical conversion of six carbon sugars into acids in bacteria
[[File:Mixed Acid Fermentation in E. coli. jpg|thumb|300px|right|The mixed acid fermentation pathway in E. coli. The mixture of end products produced by mixed acid fermentation includes lactate, acetate, succinate, formate, ethanol and the gases |H2 and |CO2. The formation of these end products depends on the presence of certain key enzymes in the bacterium. The proportion in which they are formed varies between different bacterial species. The mixed acid fermentation pathway differs from other fermentation pathways, which produce fewer end products in fixed amounts. The end products of mixed acid fermentation can have many useful applications in biotechnology and industry. For instance, ethanol is widely used as a biofuel. Therefore, multiple bacterial strains have been metabolically engineered in the laboratory to increase the individual yields of certain end products. This research has been carried out primarily in E. coli and is ongoing. Variations of mixed acid fermentation occur in a number of bacterial species, including bacterial pathogens such as Haemophilus influenzae where mostly acetate and succinate are produced and lactate can serve as a growth substrate.

Formate can be converted to hydrogen gas and carbon dioxide in E. coli. This reaction requires the enzyme formate hydrogen lyase. It can be used to prevent the conditions inside the cell becoming too acidic. If the fermentation pathway has taken place, the mixture of acids it has produced will make the solution very acidic and cause a red colour change. The methyl red test belongs to a group known as the IMViC tests.

Multiple bacterial strains have been metabolically engineered to increase the individual yields of end products formed by mixed acid fermentation. For instance, strains for the increased production of ethanol, lactate, succinate and acetate have been developed due to the usefulness of these products in biotechnology.

The E. coli strain W3110 was genetically engineered to generate 2 moles of acetate for every 1 mole of glucose that undergoes fermentation. This is known as a homoacetate pathway.

Lactate can be used to produce a bioplastic called polylactic acid (PLA). The properties of PLA depend on the ratio of the two optical isomers of lactate (D lactate and L lactate). D lactate is produced by mixed acid fermentation in E. coli. Early experiments engineered the E. coli strain RR1 to produce either one of the two optical isomers of lactate. Later experiments modified the E. coli strain KO11, originally developed to enhance ethanol production. Scientists were able to increase the yield of D lactate from fermentation by performing several deletions.

Increasing the yield of succinate from mixed acid fermentation was first done by overexpressing the enzyme PEP carboxylase. This produced a succinate yield that was approximately 3 times greater than normal. Several experiments using a similar approach have followed. Alternative approaches have altered the redox and ATP balance to optimize the succinate yield.