Secretion of recombinant proteins via the chaperone/usher pathway in Escherichia coli

: Zavialov AV, Batchikova NV, Korpela T, Petrovskaya LE, Korobko VG, Kersley J, MacIntyre S, Zav'yalov VP

Publisher: AMER SOC MICROBIOLOGY

: 2001

: APPLIED AND ENVIRONMENTAL MICROBIOLOGY

: APPL ENVIRON MICROB

: 67

: 4

: 1805

: 1814

: 10

: 0099-2240

DOI: https://doi.org/10.1128/AEM.67.4.1805-1814.2001

Fl antigen (Caf1) of Yersinia pestis is assembled via the Caf1M chaperone/Caf1A usher pathway. We investigated the ability of this assembly system to facilitate secretion of full-length heterologous proteins fused to the Caf1 subunit in Escherichia coli. Despite correct processing of a chimeric protein composed of a modified Caf1 signal peptide, mature human interleukin-1 beta (hIL-1 beta), and mature Can, the processed product (hIL-1 beta :Caf1) remained insoluble. Coexpression of this chimera with a functional Caf1M chaperone led to the accumulation of soluble hIL-1 beta :Caf1 in the periplasm. Soluble hIL-1 beta :Caf1 reacted,vith monoclonal antibodies directed against structural epitopes of hIL-1 beta. The results indicate that Caf1M-induced release of hIL-1 beta :Caf1 from the inner membrane promotes folding of the hIL-1 beta domain. Similar results were obtained with the fusion of Caf1 to hIL-1 beta receptor antagonist or to human granulocyte-macrophage colony-stimulating factor. Following coexpression of the hIL-1 beta :Caf1 precursor with both the Caf1M chaperone and Caf1A outer membrane protein, hIL-1 beta :Caf1 could he detected on the cell surface of E. coli. These results demonstrate for the first time the potential application of the chaperone/usher secretion pathway in the transport of subunits with large heterogeneous N-terminal fusions. This represents a novel means for the delivery of correctly folded heterologous proteins to the periplasm and cell surface as either polymers or cleavable monomeric domains.