Expression of catalytic subunit of bovine enterokinase in the filamentous fungus Aspergillus niger.
The cDNA encoding for catalytic subunit of bovine enterokinase (EK(L)), to which the sequence for Kex2 protease cleavage site was inserted, was expressed in the protease deficient filamentous fungus Aspergillus niger AB1.13. Fungal transformants were obtained in which expression of the glucoamylase fusion gene resulted in secretion of the protein into growth medium. Fusion polypeptide was processed to mature EK(L) by endogenous Kex-2 like protease cleavage during secretory pathway. The highest quantity of EK(L), up to 5 mg l(-1), was obtained in soya milk medium. The secreted EK(L) was easily purified from other proteins found in A. niger culture supernatant, using ion exchange and affinity chromatography. The yield of the purified and highly active EK(L) was 1.9 mg l(-1) of culture.
Svetina
M
M
National Institute of Chemistry, and Laboratory for Biosynthesis and Biotransformation, Ljubljana, Slovenia.
Krasevec
N
N
Gaberc-Porekar
V
V
Komel
R
R
D000818:Animals; D001234:Aspergillus niger / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001709:Biotechnology / Q000379:methods*; D020134:Catalytic Domain; D002417:Cattle; D002455:Cell Division; D003470:Culture Media; D004765:Enteropeptidase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D005822:Genetic Vectors; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*
D003470:Culture Media; D011994:Recombinant Proteins; D004765:Enteropeptidase