Nature Biotechnology
23, 475 - 481 (2005)
Published online: 3 April 2005; | doi:10.1038/nbt1077
Extracellular secretion of polypeptides using a modified Escherichia coli flagellar secretion apparatusKatariina Majander1, Lena Anton1, Jenni Antikainen1, Hannu Lång1, Mirko Brummer2, Timo K Korhonen1
& Benita Westerlund-Wikström11
General Microbiology, Faculty of Biosciences, P.O. Box 56, FIN-00014 University of Helsinki, Helsinki, Finland. 2
Orion Diagnostica Oy, P.O. Box 83, FIN-02101, Espoo, Finland.
Correspondence should be addressed to Benita Westerlund-Wikström benita.westerlund@helsinki.fiWe developed a modified flagellar type III secretion apparatus to secrete heterologous polypeptides into the growth medium of Escherichia coli. The secretion was facilitated by fusing the 173-bp untranslated DNA fragment upstream of the gene fliC (encoding flagellin) as well as a transcriptional terminator from fliC, into the gene encoding the polypeptide of interest. The polypeptides secreted into the growth medium at concentrations ranging from 1 to 15 mg/l were from Campylobacter jejuni (262 residues in length), Streptococcus pneumoniae (434 residues), Staphylococcus aureus (115 residues), and N-terminal FliC hybrid proteins, for example, the eukaryotic green fluorescent protein (238 residues). The expressed proteins represented >50% of total secreted protein. Previously reported protein yields from extracellular secretion of foreign proteins in E. coli have been low, approximately 100  g/l1. The strengths of our method are the concentration and purity of the secreted proteins and its versatility with regard to the proteins' length and origin.
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