| 1. |
- Gustavsson, Martin, 1984-, et al.
(author)
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Biocatalysis on the surface of Escherichia coli : melanin pigmentation of the cell exterior
- 2016
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In: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 6
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Journal article (peer-reviewed)abstract
- Today, it is considered state-of-the-art to engineer living organisms for various biotechnology applications. Even though this has led to numerous scientific breakthroughs, the enclosed interior of bacterial cells still restricts interactions with enzymes, pathways and products due to the mass-transfer barrier formed by the cell envelope. To promote accessibility, we propose engineering of biocatalytic reactions and subsequent product deposition directly on the bacterial surface. As a proof-of-concept, we used the AIDA autotransporter vehicle for Escherichia coli surface expression of tyrosinase and fully oxidized externally added tyrosine to the biopolymer melanin. This resulted in a color change and creation of a black cell exterior. The capture of ninety percent of a pharmaceutical wastewater pollutant followed by regeneration of the cell bound melanin matrix through a simple pH change, shows the superior function and facilitated processing provided by the surface methodology. The broad adsorption spectrum of melanin could also allow removal of other micropollutants.
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| 2. |
- Jarmander, Johan, et al.
(author)
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Process optimization for increased yield of surface-expressed protein in Escherichia coli
- 2014
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In: Bioprocess and biosystems engineering (Print). - : Springer Science and Business Media LLC. - 1615-7591 .- 1615-7605. ; 37:8, s. 1685-1693
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Journal article (peer-reviewed)abstract
- The autotransporter family of Gram-negative protein exporters has been exploited for surface expression of recombinant passenger proteins. While the passenger in some cases was successfully translocated, a major problem has been low levels of full-length protein on the surface due to proteolysis following export over the cytoplasmic membrane. The aim of the present study was to increase the surface expression yield of the model protein SefA, a Salmonella enterica fimbrial subunit with potential for use in vaccine applications, by reducing this proteolysis through process design using Design of Experiments methodology. Cultivation temperature and pH, hypothesized to influence periplasmic protease activity, as well as inducer concentration were the parameters selected for optimization. Through modification of these parameters, the total surface expression yield of SefA was increased by 200 %. At the same time, the yield of full-length protein was increased by 300 %, indicating a 33 % reduction in proteolysis.
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