Patent
Xylose utilization in recombinant zymomonas
| العنوان: | Xylose utilization in recombinant zymomonas |
|---|---|
| Patent Number: | 8,679,822 |
| تاريخ النشر: | March 25, 2014 |
| Appl. No: | 13/161734 |
| Application Filed: | June 16, 2011 |
| مستخلص: | Xylose-utilizing Zymomonas strains studied were found to accumulate ribulose when grown in xylose-containing media. Engineering these strains to increase ribose-5-phosphate isomerase activity led to reduced ribulose accumulation, improved growth, improved xylose utilization, and increased ethanol production. |
| Inventors: | Caimi, Perry G. (Kennett Square, PA, US); McCole, Laura (East Fallowfield, PA, US); Tao, Luan (Wallingford, PA, US); Tomb, Jean-Francois (Wilmington, DE, US); Viitanen, Paul V. (West Chester, PA, US) |
| Assignees: | E I du Pont de Nemours and Company (Wilmington, DE, US) |
| Claim: | 1. A recombinant bacterial host cell comprising: a) a xylose metabolic pathway comprising at least one gene encoding a polypeptide having xylose isomerase activity; b) at least one gene encoding a polypeptide having ribose-5-phosphate isomerase activity selected from the group consisting of ribose-5-phosphate isomerase A and ribose-5-phosphate isomerase B; and c) at least one genetic modification which increases ribose-5-phosphate isomerase activity in the host cell as compared with ribose-5-phosphate isomerase activity in the host cell lacking said genetic modification; wherein the bacterial host cell utilizes xylose to produce ethanol; and wherein the bacterial host cell is selected from the group consisting of Zymomonas and Zymobacter. |
| Claim: | 2. The recombinant host cell of claim 1 wherein the at least one genetic modification of step (c) is over-expression of an endogenous gene encoding a polypeptide having ribose-5-phosphate isomerase activity. |
| Claim: | 3. The recombinant host cell of claim 1 wherein the at least one genetic modification of step (c) is expression of at least one non-endogenous gene encoding a polypeptide having ribose-5-phosphate isomerase activity. |
| Claim: | 4. The recombinant microorganism of claim 1 wherein the polypeptide having ribose-5-phosphate isomerase activity has the EC classification EC 5.3.1.6. |
| Claim: | 5. The recombinant host cell of claim 1 wherein the ribose-5-phosphate isomerase A polypeptide: i) gives an E-value score of 0.1 or less when queried using a Profile Hidden Markov Model prepared using SEQ ID NOs:, 86, 96, and 97; the query being carried out using the hmmsearch algorithm wherein the Z parameter is set to 1 billion, and ii) has aspartic acid and glutamic acid at positions corresponding to 107 and 129, respectively, in the Saccharomyces cerevisiae RPI-A protein of SEQ ID NO:97. |
| Claim: | 6. The recombinant host cell of claim 1 wherein the ribose-5-phosphate isomerase A has an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:86, 96 and 97 based on the Clustal W method of alignment using the default parameters of GAP PENALTY=10, GAP LENGTH PENALTY=0.1, and Gonnet 250 series of protein weight matrix. |
| Claim: | 7. The recombinant host cell of claim 6 wherein the ribose-5-phosphate isomerase A has an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 86, 96, and 97 based on the Clustal W method of alignment using the default parameters of GAP PENALTY=10, GAP LENGTH PENALTY=0.1, and Gonnet 250 series of protein weight matrix. |
| Claim: | 8. The recombinant host cell of claim 1 wherein said microorganism further comprises at least one genetic modification which increases xylose isomerase activity as compared to xylose isomerase activity in the microorganism lacking said genetic modification. |
| Claim: | 9. The recombinant host cell of claim 1 wherein the gene encoding a polypeptide having xylose isomerase activity is over-expressed. |
| Claim: | 10. The recombinant microorganism of claim 1 wherein the gene encoding a polypeptide having xylose isomerase activity is expressed in multicopy. |
| Claim: | 11. A process for producing ethanol comprising: a) providing a recombinant bacterial host cell of claim 1 ; and b) culturing the bacterial host cell of (a) in a medium comprising xylose whereby xylose is converted to ethanol. |
| Claim: | 12. The process of claim 11 wherein the medium comprises either a mixture of sugars including xylose or xylose as a sole sugar. |
| Current U.S. Class: | 4352/523 |
| Patent References Cited: | 5514583 May 1996 Picataggio et al. 5712133 January 1998 Picataggio et al. 6566107 May 2003 Zhang 7223575 May 2007 Zhang et al. 7741084 June 2010 Viitanen et al. 7741119 June 2010 Viitanen et al. 7803623 September 2010 Caimi et al. 7897396 March 2011 Caimi et al. 7989206 August 2011 Viitanen et al. 7998722 August 2011 Viitanen et al. 2008/0261287 October 2008 Winkler et al. 2011/0014670 January 2011 Caimi et al. 2011/0143408 June 2011 Yang |
| Other References: | Karhumaa, Kaisa et al., Investigation of limiting metabolic steps in the utilization of xylose by recombinant Saccharomyces cervisiae using metablic engineering, Yeast, 2005, pp. 359-368, vol. 22. cited by applicant International Search Report dated Oct. 21, 2011, International Application No. PCT/US2011/042122. cited by applicant Feldmann, Sigrun D. et al., Pentose metabolism in Zymomonas mobilis wild-type and recombinant strains, Applied Microbiology and Biotechnology, 1992, pp. 354-361, vol. 38, Springer-Verlag. cited by applicant Yanase, Hideshi et al., Genetic Engineering of Zymobacter palmae for Production of Ethanol from Xylose, Applied and Environmental Microbiology, Apr. 2007, pp. 2592-2599, vol. 73, No. 8, American Society for Microbiology. cited by applicant Zhang, Min et al., Metabolic Engineering of a Pentose Metabolism Pathway in Ethanologenic Zymomonas mobilis, Science, Jan. 13, 1995, pp. 240-243, vol. 267. cited by applicant Kahsay, Robel et al., U.S. Appl. No. 13/161,749, filed Jun. 16, 2011. cited by applicant |
| Assistant Examiner: | Chowdhury, Iqbal H |
| Primary Examiner: | Mondesi, Robert |
| رقم الانضمام: | edspgr.08679822 |
| قاعدة البيانات: | USPTO Patent Grants |
| الوصف غير متاح. |