Forward genetics screen coupled with whole-genome resequencing identifies novel gene targets for improving heterologous enzyme production in Aspergillus niger
Abstract
Plant biomass, once reduced to its composite sugars, can be converted to fuel substitutes. One means of overcoming the recalcitrance of lignocellulose is pretreatment followed by enzymatic hydrolysis. However, currently available commercial enzyme cocktails are inhibited in the presence of residual pretreatment chemicals. Recent studies have identified a number of cellulolytic enzymes from bacteria that are tolerant to pretreatment chemicals such as ionic liquids. The challenge now is generation of these enzymes in copious amounts, an arena where fungal organisms such as Aspergillus niger have proven efficient. Fungal host strains still need to be engineered to increase production titers of heterologous protein over native enzymes, which has been a difficult task. Here, we developed a forward genetics screen coupled with whole-genome resequencing to identify specific lesions responsible for a protein hyper-production phenotype in A. niger. This strategy successfully identified novel targets, including a low-affinity glucose transporter, MstC, whose deletion significantly improved secretion of recombinant proteins driven by a glucoamylase promoter.
- Authors:
-
[1]
- Joint BioEnergy Institute, Emeryville, CA (United States); Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
- Joint BioEnergy Institute, Emeryville, CA (United States); Wright-Patterson Air Force Base, Dayton, OH (United States)
- Joint BioEnergy Institute, Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
- Joint BioEnergy Institute, Emeryville, CA (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States)
- Publication Date:
- Research Org.:
- Pacific Northwest National Laboratory (PNNL), Richland, WA (United States); Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1416031
- Alternate Identifier(s):
- OSTI ID: 1420441; OSTI ID: 1431407; OSTI ID: 1433118
- Report Number(s):
- PNNL-SA-132880
Journal ID: ISSN 0175-7598; PII: 8717
- Grant/Contract Number:
- AC02-05CH11231; AC05-76RL01830
- Resource Type:
- Published Article
- Journal Name:
- Applied Microbiology and Biotechnology
- Additional Journal Information:
- Journal Name: Applied Microbiology and Biotechnology Journal Volume: 102 Journal Issue: 4; Journal ID: ISSN 0175-7598
- Publisher:
- Springer
- Country of Publication:
- Germany
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; Aspergillus niger; Heterologous protein expression; Forward mutagenesis; Whole-genome resequencing; MstC
Citation Formats
Reilly, Morgann C., Kim, Joonhoon, Lynn, Jed, Simmons, Blake A., Gladden, John M., Magnuson, Jon K., and Baker, Scott E. Forward genetics screen coupled with whole-genome resequencing identifies novel gene targets for improving heterologous enzyme production in Aspergillus niger. Germany: N. p., 2018.
Web. doi:10.1007/s00253-017-8717-3.
Reilly, Morgann C., Kim, Joonhoon, Lynn, Jed, Simmons, Blake A., Gladden, John M., Magnuson, Jon K., & Baker, Scott E. Forward genetics screen coupled with whole-genome resequencing identifies novel gene targets for improving heterologous enzyme production in Aspergillus niger. Germany. https://doi.org/10.1007/s00253-017-8717-3
Reilly, Morgann C., Kim, Joonhoon, Lynn, Jed, Simmons, Blake A., Gladden, John M., Magnuson, Jon K., and Baker, Scott E. Sat .
"Forward genetics screen coupled with whole-genome resequencing identifies novel gene targets for improving heterologous enzyme production in Aspergillus niger". Germany. https://doi.org/10.1007/s00253-017-8717-3.
@article{osti_1416031,
title = {Forward genetics screen coupled with whole-genome resequencing identifies novel gene targets for improving heterologous enzyme production in Aspergillus niger},
author = {Reilly, Morgann C. and Kim, Joonhoon and Lynn, Jed and Simmons, Blake A. and Gladden, John M. and Magnuson, Jon K. and Baker, Scott E.},
abstractNote = {Plant biomass, once reduced to its composite sugars, can be converted to fuel substitutes. One means of overcoming the recalcitrance of lignocellulose is pretreatment followed by enzymatic hydrolysis. However, currently available commercial enzyme cocktails are inhibited in the presence of residual pretreatment chemicals. Recent studies have identified a number of cellulolytic enzymes from bacteria that are tolerant to pretreatment chemicals such as ionic liquids. The challenge now is generation of these enzymes in copious amounts, an arena where fungal organisms such as Aspergillus niger have proven efficient. Fungal host strains still need to be engineered to increase production titers of heterologous protein over native enzymes, which has been a difficult task. Here, we developed a forward genetics screen coupled with whole-genome resequencing to identify specific lesions responsible for a protein hyper-production phenotype in A. niger. This strategy successfully identified novel targets, including a low-affinity glucose transporter, MstC, whose deletion significantly improved secretion of recombinant proteins driven by a glucoamylase promoter.},
doi = {10.1007/s00253-017-8717-3},
journal = {Applied Microbiology and Biotechnology},
number = 4,
volume = 102,
place = {Germany},
year = {Sat Jan 06 00:00:00 EST 2018},
month = {Sat Jan 06 00:00:00 EST 2018}
}
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https://doi.org/10.1007/s00253-017-8717-3
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Cited by: 11 works
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Figures / Tables:
Table 1: Strains used in this study
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