Award Abstract # 1456071
SBIR Phase II: Large-scale, high-throughput optimization of gene expression in industrial yeast for improved small molecule production

NSF Org: TI
Translational Impacts
Recipient: LYGOS, INC.
Initial Amendment Date: February 18, 2015
Latest Amendment Date: May 26, 2020
Award Number: 1456071
Award Instrument: Standard Grant
Program Manager: Erik Pierstorff
epiersto@nsf.gov
 (703)292-0000
TI
 Translational Impacts
TIP
 Directorate for Technology, Innovation, and Partnerships
Start Date: March 1, 2015
End Date: October 31, 2020 (Estimated)
Total Intended Award Amount: $750,000.00
Total Awarded Amount to Date: $1,425,979.00
Funds Obligated to Date: FY 2015 = $750,000.00
FY 2016 = $159,979.00

FY 2017 = $16,000.00

FY 2018 = $500,000.00
History of Investigator:
  • Andrew Conley (Principal Investigator)
    aconley@lygos.com
  • Jeffrey Dietrich (Former Principal Investigator)
Recipient Sponsored Research Office: Lygos Inc.
25821 INDUSTRIAL BLVD STE 300
HAYWARD
CA  US  94545-2919
(605)679-7717
Sponsor Congressional District: 14
Primary Place of Performance: Lygos, Inc
5858 Horton St., Suite 410
Emeryville
CA  US  94608-2006
Primary Place of Performance
Congressional District:
12
Unique Entity Identifier (UEI): FR5ECFM6UKL6
Parent UEI:
NSF Program(s): SBIR Phase II
Primary Program Source: 01001516DB NSF RESEARCH & RELATED ACTIVIT
01001617DB NSF RESEARCH & RELATED ACTIVIT

01001718DB NSF RESEARCH & RELATED ACTIVIT

01001819DB NSF RESEARCH & RELATED ACTIVIT
Program Reference Code(s): 116E, 165E, 169E, 5373, 8038, 8240, 9102, 9231, 9251
Program Element Code(s): 537300
Award Agency Code: 4900
Fund Agency Code: 4900
Assistance Listing Number(s): 47.084

ABSTRACT

The broader impact/commercial potential of this Small Business Innovation Research (SBIR) Phase II project is development of a microbial technology for the conversion of low-value sugars into high-value chemicals. Most industrial chemicals produced today are derived from petroleum and other nonrenewable raw materials. The long-term growth and sustainability of the chemical industry benefits from development of new routes to existing chemicals using renewable raw materials. Furthermore, due to higher infrastructure costs and stricter environmental requirements, many chemicals that were once produced in the United States are now produced abroad. This contributes to the U.S. trade deficit. This Phase II proposal aims to develop a fermentation technology where domestically grown agricultural materials (for example, corn and waste agricultural residues) are converted into high-value chemicals. The optimized fermentation process is estimated to be cost-competitive with the incumbent petrochemical route when scaled. If successful, this proposal will facilitate growth of a domestic bio-chemical manufacturing industry, targeting the $30 billion organic acids market.

This SBIR Phase II project proposes to develop large-scale, high-throughput techniques to optimize gene expression in industrial yeast. A significant problem within the field of industrial biotechnology is the ability to engineer and optimize the fermentation performance of non-academic or model microbes. Most molecular metabolic engineering tools are developed for use in two model prokaryotic and eukaryotic microbes, E. coli and S. cerevisiae, and are not suitable for use with industrially relevant microbes. Without these tools it is costly and slow to commercialize new fermentation technologies. The goal of this Phase II project is to develop and implement a set of molecular biology tools designed for acid-tolerant yeast, and working to apply them toward improving small molecule production. Specifically, the molecular biology tools are useful for tuning (up- or down-regulation) user-defined gene transcription and translation. Engineered microbes harboring the desired genetic modification(s) are assayed for improved small molecule production from sugar in small scale fermentations. Successful genetic modifications are those that result in more efficient small molecule product formation from sugar, and ideally decreased biomass formation from sugar, providing a lower production cost in a scaled, commercial process.

PROJECT OUTCOMES REPORT

Disclaimer

This Project Outcomes Report for the General Public is displayed verbatim as submitted by the Principal Investigator (PI) for this award. Any opinions, findings, and conclusions or recommendations expressed in this Report are those of the PI and do not necessarily reflect the views of the National Science Foundation; NSF has not approved or endorsed its content.

Lygos has developed an engineered yeast that converts glucose to malonic acid, a high-value chemical with diverse applications as an intermediate chemical across several industries today, including flavors and fragrances, pharmaceuticals, semiconductor manufacturing and bio-polymers. Lygos scientists and engineers are currently working on developing a second generation (Gen2) strain and accompanying fermentation process in order to capture additional, incremental improvements in performance. This will decrease production costs to enable new application development and entry into larger-volume, lower-price point markets.

In more detail, this NSF-funded project is directed toward improving a key strain performance metric, specific productivity (Qp; the rate of product formed per amount of biocatalyst dry cell weight; g-malonic acid/g-DCW/hr), which we have determined has the most significant impact on commercial production costs. Based on our internal analysis of intermediate and byproduct accumulation in our fermentations, we concluded that acetyl-CoA carboxylase (ACC) was the rate-limiting enzyme in the pathway. Thus, the goal of this project was to identify a strain with enhanced acetyl-CoA carboxylase activity by testing various factors believed to have an impact on ACC activity. We screened combinatorial libraries of engineered strain variants using our proprietary malonic acid yeast strain. Factors evaluated included, but were not limited to testing the following: varying expression levels of the ACC, expressing heterologous ACC homologs, improving enzyme cofactor availability, mitigating negative enzyme regulatory effects, altering expression of the other malonic acid pathway enzymes, enhancing the strain's ability to export malonate and augmenting fatty acid biosynthesis. As well, CO2 supplementation studies were investigated to optimize the fermentation process.

Over 2,000 unique strains were generated during this project, leading to strains with significantly improved Qp values that surpassed all Milestone objectives for this grant, resulting in improved production strains for malonic acid.

 


Last Modified: 02/03/2021
Modified by: Andrew Conley

Please report errors in award information by writing to: awardsearch@nsf.gov.

Print this page

Back to Top of page