High-level expression of the thermoalkalophilic lipase from Bacillus thermocatenulatus in Escherichia coli

M. L. Rúa, H. Atomi, C. Schmidt-Dannert, R. D. Schmid

Research output: Contribution to journalArticlepeer-review

43 Scopus citations


An efficient expression system for the previously only weakly expressed thermophilic lipase BTL2 (Bacillus thermocatenulatus lipase 2) was developed for the production of large amounts of lipase in Escherichia coli. Therefore, the gene was subcloned in the pCYT-EXP1 (pT1) expression vector downstream of the temperature-inducible λ promoter P(L). Three different expression vectors were constructed: (i) pT1-BTL2 containing the mature lipase gene, (ii) pT1-preBTL2 containing the prelipase gene and (iii) pT1-OmpABTL2 containing the mature lipase gene fused to the signal peptide of the OmpA protein, the major outer membrane protein of E. coli. With pT1-BTL2 and pT1-preBTL2, comparable expression levels of 7000-9000 U/g cells were obtained independently of the E. coli host. In contrast, with E. coli JM105 harbouring pT1-OmpABTL2, 660 000 soluble lipase U/g cells was produced, whereas, with E. coli JM105 and BL321, production levels of 30 000 U/g cells were achieved. However, most of the lipase remained insoluble but active after cell breakage because of the unprocessed OmpA signal peptide. A simple cholate extraction followed by proteinase K cleavage and ultrafiltration allowed the isolation of 1.15 x 106 units of 90% pure mature lipase/wet cells.

Original languageEnglish (US)
Pages (from-to)405-410
Number of pages6
JournalApplied Microbiology and Biotechnology
Issue number4
StatePublished - 1998

Bibliographical note

Funding Information:
Acknowledgements Dr. M. Luisa Rúa gratefully acknowledges a scholarship from the Spanish Ministry of Education and Science. Dr. H. Atomi gratefully acknowledges a scholarship from the CIBA-GEIGY Foundation (Japan). We would like to thank Mrs. R. Getzlaff for N-terminal sequencing.


Dive into the research topics of 'High-level expression of the thermoalkalophilic lipase from Bacillus thermocatenulatus in Escherichia coli'. Together they form a unique fingerprint.

Cite this