Unconventional protein secretion
AssigneeHeinrich-Heine-Universitaet Duesseldorf
InventorMichael Feldbruegge
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Abstract
The present invention relates to an expression cassette comprising a nucleotide sequence encoding an amino acid sequence, a fragment or variant thereof which directs unconventional protein secretion and a nucleotide sequence encoding a protein of interest. Also contemplated is a vector which comprises the expression cassette, host cells comprising the vectors as well as methods and uses for the production of a polypeptide of interest.
Claims25
An expression cassette comprising
- (a) a nucleotide sequence encoding
- (i) an amino acid sequence having amino acids n-502 of the amino acid sequence shown in SEQ ID No:2, wherein n is amino acid position 1 of SEQ ID No:2, or a fragment thereof which directs unconventional protein secretion, or
- (ii) an amino acid sequence which is at least 60% identical to the amino acid sequence of (i) and which directs unconventional protein secretion;
- and
- (b) a nucleotide sequence encoding a protein of interest,
- wherein nucleotide sequence (a) and (b) are fused in frame,
- with the proviso that nucleotide sequence (b) does not encode green fluorescence protein.
10-11. (canceled)
13-20. (canceled)
22-26. (canceled)
29-31. (canceled)
A method for identifying an amino acid sequence which directs unconventional protein secretion, comprising
- (a) providing a host cell expressing a fusion protein comprising (i) an amino acid sequence which is suspected to direct unconventional protein secretion and (ii) an amino acid sequence encoding a marker protein having a detectable activity which is subject to N-glycosylation in said host cell, thereby inactivating said marker protein,
- (b) determining whether said marker protein is secreted by said host cell by detecting its activity,
- wherein said amino acid sequence which is suspected to direct unconventional protein secretion directs unconventional protein secretion if it is active after secretion by said host cell.
(canceled)
Use of β-glucuronidase (Gus) for the identification of an amino acid sequence which directs unconventional protein secretion.
Dependencies are inferred from the claim text.