Methods of increasing secretion of polypeptides having biological activity
The present invention relates to methods for producing a secreted polypeptide having biological activity, comprising: (a) transforming a fungal host cell with a fusion protein construct encoding a fusion protein, which comprises: (i) a first polynucleotide encoding a signal peptide; (ii) a second polynucleotide encoding at least a catalytic domain of an endoglucanase or a portion thereof; and (iii) a third polynucleotide encoding at least a catalytic domain of a polypeptide having biological activity; wherein the signal peptide and at least the catalytic domain of the endoglucanase increases secretion of the polypeptide having biological activity compared to the absence of at least the catalytic domain of the endoglucanase; (b) cultivating the transformed fungal host cell under conditions suitable for production of the fusion protein; and (c) recovering the fusion protein, a component thereof, or a combination thereof, having biological activity, from the cultivation medium.
1. An isolated fusion protein, comprising:
(a) a first amino acid sequence comprising a signal peptide;
(b) a second amino acid sequence comprising at least a catalytic domain of a Cel45 endoglucanase; and
(c) a third amino acid sequence comprising at least a catalytic domain of a polypeptide having biological activity, wherein said fusion protein comprises SEQ ID NO: 74 or SEQ ID NO: 76.
2. The fusion protein of claim 1 , which further comprises a carbohydrate binding module.
3. The fusion protein of claim 1 , which further comprises a linker located between the endoglucanase catalytic domain and the catalytic domain of the polypeptide having biological activity and a cleavage site located between the endoglucanase catalytic domain and/or the catalytic domain of a polypeptide having biological activity.
4. The fusion protein of claim 1 , which further comprises a fourth amino acid sequence comprising a second signal peptide.
5. The fusion protein of claim 1 , wherein the second signal peptide is linked in frame to the N-terminus of the amino acid sequence comprising the catalytic domain of the polypeptide having biological activity or at least the catalytic domain of the endoglucanase.
6. The fusion protein of claim 1 , wherein SEQ ID NO: 74 is encoded by SEQ ID NO: 73 and SEQ ID NO: 76 is encoded by SEQ ID NO: 75.