Recombinant Zymomonas for pentose fermentation

• US 5,726,053 A
• Filed: 04/14/1995
• Issued: 03/10/1998
• Est. Priority Date: 04/15/1994
• Status: Expired due to Term

• Current Assignee: Alliance For Sustainable Energy LLC
• Original Assignee: Midwest Research Institute
• Inventors: Zhang, Min, Picataggio, Stephen K., Eddy, Christina K., Deanda, Kristine A.
• Primary Examiner(s): Prouty, Rebecca E.
• Application Number:
  US08/421,996
• Time in Patent Office: 1,061 Days
• Field of Search: 435/252.3, 435/243, 435/320.1, 435/161, 435/163, 435/165, 435/822, 536/23.2, 536/23.7, 935/29, 935/60, 935/72
• US Class Current: 435/252.3
• CPC Class Codes: C12N 15/52 Genes encoding for enzymes or proenzymes
• CPC Class Codes: C12N 9/10 Transferases (2.) ribonucleases C12N9/22
• CPC Class Codes: C12N 9/90 Isomerases (5.)
• CPC Class Codes: C12P 7/065 with microorganisms other than yeasts
• CPC Class Codes: Y02E 50/10 Biofuels, e.g. bio-diesel
• CPC Class Codes: Y10S 435/822 using bacteria or actinomycetales

Abstract

The invention relates to microorganisms which normally do not ferment pentose sugar and which are genetically altered to ferment pentose sugar to produce ethanol, and fermentation processes utilizing the same. Examples include Zymomonas mobilis which has been transformed with combinations of E. coli genes for xylose isomerase, xylulokinase, transaldolase, transketolase, L-arabinose isomerase, L-ribulokinase, and L-ribulose-5-phosphate 4-epimerase. Expression of the added genes are under the control of Zymomonas mobilis promoters. These newly created microorganisms are useful for fermenting pentoses and glucose, produced by hydrolysis of hemicellulose and cellulose, to produce ethanol.

Claims

1. The microorganism Zymomonas mobilis containing exogenous genes encoding L-arabinose isomerase, L-ribulokinase, L-ribulose-5-phosphate-4-epimerase, transaldolase and transketolase which is capable of growing on arabinose as a sole carbon source and fermenting said arabinose to ethanol, wherein said microorganism without said genes is incapable of fermenting said arabinose to ethanol.

2. The microorganism of claim 1, wherein said transaldolase and transketolase genes, are expressed under the control of the Z. mobilis enolase promoter.

3. The microorganism of claim 1, wherein said

L-arabinose isomerase, L-ribulokinase, and L-ribulose 5-phosphate 4-epimerase genes are expressed under the control Z. mobilis glyceraldehyde-3-phosphate dehydrogenase promotor.

4. The microorganism of claim 1 wherein said exogenous genes are obtained from a microorganism selected from the group consisting of E. coli, Xanthomonas, Klebsiella, Rhodobacter, Flavobacterium, Acetobacter, Gluconobacter, Rhizobium, Agrobacterium, Salmonella and Pseudomonads.

5. The microorganism of claim 1 wherein said exogenous genes are expressed under the control of one or more promoters located 5'"'"' to said genes, whereby said genes are expressed in said microorganism.

6. The microorganism of claim 4, wherein the genes are integrated into the host genome.

7. The microorganism of claim 4, wherein said genes are contained on a vector.

8. The microorganism of claim 7, wherein said vector is a plasmid.

10. The microorganism of claim 1, which contains a vector comprising genes encoding L-arabinose isomerase, L-ribulokinase, L-ribulose 5-phosphate 4-epimerase, transaldolase and transketolase, and at least one promoter selected from the group consisting of Z. mobilis glyceraldehyde-3-phosphate dehydrogenase and Z. mobilis enolase recognized by Z. mobilis which regulates the expression of the genes.

11. The microorganism of claim 4, containing exogenous genes which encode L-arabinose isomerase, L-ribulokinase, and L-ribulose 5-phosphate 4-epimerase, expressed under the control of the Z. mobilis glyceraldehyde-3-phosphate dehydrogenase promoter, and exogenous genes which encode transaldolase and transketolase, expressed under the control of the Z. mobilis enolase promoter.

12. The microorganism of claim 1, wherein, said genes are coordinately expressed.

9. A vector for the transformation of Zymomonas mobilis comprising genes encoding L-arabinose isomerase, L-ribulokinase, L-ribulose 5-phosphate 4-epimerase, transaldolase and transketolase, and at least one promotor selected from the group consisting of Z. mobilis glyceraldehyde-3-phosphate dehydrogenase and Z. mobilis enolase recognized by Zymomonas mobilis which regulates the expression of the genes.