Surface Expression of omega-Transaminase in Escherichia coli
2014 (English)In: Applied and Environmental Microbiology, ISSN 0099-2240, E-ISSN 1098-5336, Vol. 80, no 7, 2293-2298 p.Article in journal (Refereed) Published
Chiral amines are important for the chemical and pharmaceutical industries, and there is rapidly growing interest to use transaminases for their synthesis. Since the cost of the enzyme is an important factor for process economy, the use of whole-cell biocatalysts is attractive, since expensive purification and immobilization steps can be avoided. Display of the protein on the cell surface provides a possible way to reduce the mass transfer limitations of such biocatalysts. However, transaminases need to dimerize in order to become active, and furthermore, they require the cofactor pyridoxal phosphate; consequently, successful transaminase surface expression has not been reported thus far. In this work, we produced an Arthrobacter citreus omega-transaminase in Escherichia coli using a surface display vector based on the autotransporter adhesin involved in diffuse adherence (AIDA-I), which has previously been used for display of dimeric proteins. The correct localization of the transaminase in the E. coli outer membrane and its orientation toward the cell exterior were verified. Furthermore, transaminase activity was detected exclusively in the outer membrane protein fraction, showing that successful dimerization had occurred. The transaminase was found to be present in both full-length and proteolytically degraded forms. The removal of this proteolysis is considered to be the main obstacle to achieving sufficient whole-cell transaminase activity.
Place, publisher, year, edition, pages
2014. Vol. 80, no 7, 2293-2298 p.
Display, Protein, Autodisplay, Amines
Biocatalysis and Enzyme Technology
IdentifiersURN: urn:nbn:se:kth:diva-122223DOI: 10.1128/AEM.03678-13ISI: 000332840700028ScopusID: 2-s2.0-84896029013OAI: oai:DiVA.org:kth-122223DiVA: diva2:621413
QC 20140423. Updated from submitted to published.2013-05-142013-05-142014-04-23Bibliographically approved