Abstract
The gene encoding the aspartic protease in the mold Mucor miehei was isolated and a vector, pMA67, was constructed to allow expression of this gene in Mucor circinelloides. pMA67 contained the Amp gene for selection in E. coli, a Leu+ gene for selection in M. circinelloides and the M. miehei aspartic prepro-protease gene. A leucine requiring strain of M. circinelloides was transformed with pMA67. It was shown by rocket immunoelectrophoresis that transformants secreted M. miehei aspartic protease. The excreted protease is active and has the same apparent molecular weight as the aspartic protease produced by M. miehei. This paper reports on heterologous gene expression and protein secretion in a member of the commercially important genus Mucor, and it opens up the possibility to use pMA67 for the expression of other important proteins.
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Abbreviations
- Amp:
-
ampicillin
- bp:
-
base pair(s)
- EDTA:
-
ethylene diamine tetraacetic acid, sodium salt
- kb:
-
1000 bp
- IPTG:
-
isopropyl β-d-thiogalactoside
- McAP:
-
Mucor circinelloides aspartic protease
- MmAP:
-
Mucor miehei aspartic protease
- SDS:
-
sodium dodecyl sulphate
- TRIS:
-
tris(hydroxymethyl)-amino methane
- X-gal:
-
5-bromo 4-chloro-3-indolyl-β-d-galactopyranoside
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Dickinson, L., Harboe, M., van Heeswijck, R. et al. Expression of active mucor miehei aspartic protease in Mucor circinelloides. Carlsberg Res. Commun. 52, 243 (1987). https://doi.org/10.1007/BF02907167
DOI: https://doi.org/10.1007/BF02907167