Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes
Abstract Background Heterologous production of cold-adapted proteins currently represents one of the greatest bottlenecks in the ongoing bioprospecting efforts to find new enzymes from low-temperature environments, such as, the polar oceans that represent essentially untapped resources in this respe...
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ftdoajarticles:oai:doaj.org/article:a285bccb42fa43b6bc577425b03af432 2023-05-15T15:14:10+02:00 Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes Jenny Johansson Söderberg Miriam Grgic Erik Hjerde Peik Haugen 2019-11-01T00:00:00Z https://doi.org/10.1186/s12934-019-1247-1 https://doaj.org/article/a285bccb42fa43b6bc577425b03af432 EN eng BMC http://link.springer.com/article/10.1186/s12934-019-1247-1 https://doaj.org/toc/1475-2859 doi:10.1186/s12934-019-1247-1 1475-2859 https://doaj.org/article/a285bccb42fa43b6bc577425b03af432 Microbial Cell Factories, Vol 18, Iss 1, Pp 1-16 (2019) Aliivibrio wodanis Heterologous protein production Expression host Psychrophilic enzymes Cold adaptation Marine biotechnology Microbiology QR1-502 article 2019 ftdoajarticles https://doi.org/10.1186/s12934-019-1247-1 2022-12-31T01:59:50Z Abstract Background Heterologous production of cold-adapted proteins currently represents one of the greatest bottlenecks in the ongoing bioprospecting efforts to find new enzymes from low-temperature environments, such as, the polar oceans that represent essentially untapped resources in this respect. In mesophilic expression hosts such as Escherichia coli, cold-adapted enzymes often form inactive aggregates. Therefore it is necessary to develop new low-temperature expression systems, including identification of new host organisms and complementary genetic tools. Psychrophilic bacteria, including Pseudoalteromonas haloplanktis, Shewanella and Rhodococcus erythropolis have all been explored as candidates for such applications. However to date none of these have found widespread use as efficient expression systems, or are commercially available. In the present work we explored the use of the sub-Arctic bacterium Aliivibrio wodanis as a potential host for heterologous expression of cold-active enzymes. Results We tested 12 bacterial strains, as well as available vectors, promoters and reporter systems. We used RNA-sequencing to determine the most highly expressed genes and their intrinsic promoters in A. wodanis. In addition we examined a novel 5′-fusion to stimulate protein production and solubility. Finally we tested production of a set of “difficult-to-produce” enzymes originating from various bacteria and one Archaea. Our results show that cold-adapted enzymes can be produced in soluble and active form, even in cases when protein production failed in E. coli due to the formation of inclusion bodies. Moreover, we identified a 60-bp/20-aa fragment from the 5′-end of the AW0309160_00174 gene that stimulates expression of Green Fluorescent Protein and improves production of cold-active enzymes when used as a 5′-fusion. A 25-aa peptide from the same protein enhanced secretion of a 25-aa-sfGFP fusion. Conclusions Our results indicate the use of A. wodanis and associated genetic tools for low-temperature protein ... Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic Microbial Cell Factories 18 1 |
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op_collection_id |
ftdoajarticles |
language |
English |
topic |
Aliivibrio wodanis Heterologous protein production Expression host Psychrophilic enzymes Cold adaptation Marine biotechnology Microbiology QR1-502 |
spellingShingle |
Aliivibrio wodanis Heterologous protein production Expression host Psychrophilic enzymes Cold adaptation Marine biotechnology Microbiology QR1-502 Jenny Johansson Söderberg Miriam Grgic Erik Hjerde Peik Haugen Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
topic_facet |
Aliivibrio wodanis Heterologous protein production Expression host Psychrophilic enzymes Cold adaptation Marine biotechnology Microbiology QR1-502 |
description |
Abstract Background Heterologous production of cold-adapted proteins currently represents one of the greatest bottlenecks in the ongoing bioprospecting efforts to find new enzymes from low-temperature environments, such as, the polar oceans that represent essentially untapped resources in this respect. In mesophilic expression hosts such as Escherichia coli, cold-adapted enzymes often form inactive aggregates. Therefore it is necessary to develop new low-temperature expression systems, including identification of new host organisms and complementary genetic tools. Psychrophilic bacteria, including Pseudoalteromonas haloplanktis, Shewanella and Rhodococcus erythropolis have all been explored as candidates for such applications. However to date none of these have found widespread use as efficient expression systems, or are commercially available. In the present work we explored the use of the sub-Arctic bacterium Aliivibrio wodanis as a potential host for heterologous expression of cold-active enzymes. Results We tested 12 bacterial strains, as well as available vectors, promoters and reporter systems. We used RNA-sequencing to determine the most highly expressed genes and their intrinsic promoters in A. wodanis. In addition we examined a novel 5′-fusion to stimulate protein production and solubility. Finally we tested production of a set of “difficult-to-produce” enzymes originating from various bacteria and one Archaea. Our results show that cold-adapted enzymes can be produced in soluble and active form, even in cases when protein production failed in E. coli due to the formation of inclusion bodies. Moreover, we identified a 60-bp/20-aa fragment from the 5′-end of the AW0309160_00174 gene that stimulates expression of Green Fluorescent Protein and improves production of cold-active enzymes when used as a 5′-fusion. A 25-aa peptide from the same protein enhanced secretion of a 25-aa-sfGFP fusion. Conclusions Our results indicate the use of A. wodanis and associated genetic tools for low-temperature protein ... |
format |
Article in Journal/Newspaper |
author |
Jenny Johansson Söderberg Miriam Grgic Erik Hjerde Peik Haugen |
author_facet |
Jenny Johansson Söderberg Miriam Grgic Erik Hjerde Peik Haugen |
author_sort |
Jenny Johansson Söderberg |
title |
Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
title_short |
Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
title_full |
Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
title_fullStr |
Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
title_full_unstemmed |
Aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
title_sort |
aliivibrio wodanis as a production host: development of genetic tools for expression of cold-active enzymes |
publisher |
BMC |
publishDate |
2019 |
url |
https://doi.org/10.1186/s12934-019-1247-1 https://doaj.org/article/a285bccb42fa43b6bc577425b03af432 |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
Microbial Cell Factories, Vol 18, Iss 1, Pp 1-16 (2019) |
op_relation |
http://link.springer.com/article/10.1186/s12934-019-1247-1 https://doaj.org/toc/1475-2859 doi:10.1186/s12934-019-1247-1 1475-2859 https://doaj.org/article/a285bccb42fa43b6bc577425b03af432 |
op_doi |
https://doi.org/10.1186/s12934-019-1247-1 |
container_title |
Microbial Cell Factories |
container_volume |
18 |
container_issue |
1 |
_version_ |
1766344649179398144 |