Process optimization for increased yield of surface-expressed protein in Escherichia coli
2014 (English)In: Bioprocess and biosystems engineering (Print), ISSN 1615-7591, E-ISSN 1615-7605, Vol. 37, no 8, 1685-1693 p.Article in journal (Refereed) Published
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.
Place, publisher, year, edition, pages
2014. Vol. 37, no 8, 1685-1693 p.
Surface expression, Autotransport, Process optimization, Proteolysis, Live vaccines
IdentifiersURN: urn:nbn:se:kth:diva-149969DOI: 10.1007/s00449-014-1141-5ISI: 000339962400023ScopusID: 2-s2.0-84925884851OAI: oai:DiVA.org:kth-149969DiVA: diva2:743511
QC 201409042014-09-042014-08-292015-05-08Bibliographically approved