Microorganisms having enhanced resistance to acetate and methods of use
The present invention provides isolated or genetically modified strains of microorganisms that display enhanced resistance to acetate as a result of increased expression of a sodium proton antiporter. The present invention also provides methods for producing such microbial strains, as well as related promoter sequences and expression vectors. Further, the present invention provides methods of producing alcohol from biomass materials by using microorganisms with enhanced resistance to acetate.
1. A method of producing ethanol from a cellulosic biomass material, comprising:
(a) providing a microorganism genetically modified to contain an expression vector coding for a sodium-proton antiporter, wherein the genetic modification results in increased expression of a sodium-proton antiporter in said microorganism and elevated resistance to acetate as compared to an unmodified strain of said microorganism;
(b) adding the genetically modified microorganism of (a) to a fermentation mixture comprising a cellulosic biomass material and/or fermentation substrates derived from said cellulosic biomass material;
(c) allowing said microorganism to ferment and produce ethanol; and
(d) recovering ethanol produced.
2. The method of claim 1 , wherein said microorganism is selected from bacteria or fungi.
3. The method of claim 2 , wherein said microorganism is a bacterial species selected from Acetobacterium, Bacillus, Streptococcus, Clostridium, Zymomonas sp., and Gluconobacter sp.
4. The method of claim 2 , wherein said microorganism is a fungal species selected from Saccharomyces sp., Kluyveromyces sp., Pichia sp., Candida sp., and Schizosaccharomycetes sp.
5. The method of claim 1 , wherein said sodium-proton antiporter is selected from the group consisting of a plasma membrane sodium-proton antiporter, an endosomal sodium-proton antiporter, and a vacuolar sodium-proton antiporter.
6. The method of claim 5 , wherein said sodium-proton antiporter is a plasma membrane sodium-proton antiporter native to said microorganism.
7. The method of claim 1 , wherein said microorganism is Z. mobilis.
8. The method of claim 7 , wherein said sodium-proton antiporter comprises the protein sequence as set forth in SEQ ID NO: 2.
9. The method of claim 1 , wherein said microorganism is S. cerevisiae.
10. The method of claim 9 , wherein said sodium-proton antiporter is the plasma membrane sodium-proton antiporter comprising the amino acid sequence of SEQ ID NO: 15.
11. The method of claim 1 , wherein the expression vector is an integrative vector.
12. The method of claim 1 , wherein the expression vector is a replicative vector.