Methods for the production and secretion of modified peptides
First Claim
1. A nucleic acid encoding a synthetic polypeptide, the polypeptide comprising:
- a first, non-lantibiotic export signal peptide that is SEQ ID NO;
46 or a signal for the secretory (Sec) system from Lactococcus lactis;
a second, lantibiotic leader peptide that is recognized by a lantibiotic dehydratase selected from the group consisting of lantibiotic dehydratase (LanB) and lantibiotic bifunctional enzyme (LanM) dehydratase; and
a third peptide of interest comprising at least one serine or threonine residue that can be posttranslationally dehydrated by LanB, wherein the peptide of interest is positioned C-terminally of both the non-lantibiotic export signal peptide and the lantibiotic leader peptide.
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Abstract
The invention relates to the production of heterologous polypeptides in a recombinant host cell. More specifically, it relates to the production and secretion of peptides, such as biologically active peptides, that are modified by one or more lantibiotic-synthesizing enzymes. Provided is a nucleic acid construct encoding a polypeptide comprising 1) a non-lantibiotic export signal that is recognized by a non-lantibiotic export system; 2) a lantibiotic leader peptide that is recognized by at least a lantibiotic dehydratase such as LanB and, C-terminally of said export signal and said leader peptide, 3) a peptide of interest containing one or more serine or threonine residue(s) which can be posttranslationally dehydrated by said dehydratase. Also provided is a method for producing a polypeptide in a host cell, comprising providing a host cell with a nucleic acid construct according to the invention and allowing expression and secretion of the encoded polypeptide by said host cell, wherein said host cell comprises at least a dehydratase capable of modifying said encoded polypeptide and a non-lantibiotic export system capable of secreting said modified polypeptide.
4 Citations
22 Claims
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1. A nucleic acid encoding a synthetic polypeptide, the polypeptide comprising:
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a first, non-lantibiotic export signal peptide that is SEQ ID NO;
46 or a signal for the secretory (Sec) system from Lactococcus lactis;
a second, lantibiotic leader peptide that is recognized by a lantibiotic dehydratase selected from the group consisting of lantibiotic dehydratase (LanB) and lantibiotic bifunctional enzyme (LanM) dehydratase; and a third peptide of interest comprising at least one serine or threonine residue that can be posttranslationally dehydrated by LanB, wherein the peptide of interest is positioned C-terminally of both the non-lantibiotic export signal peptide and the lantibiotic leader peptide. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)
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15. A nucleic acid encoding a synthetic polypeptide, the polypeptide comprising:
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a first peptide consisting of SEQ ID NO;
46;a second, lantibiotic leader peptide; and a third peptide of interest comprising at least one serine or threonine residue that can be posttranslationally dehydrated by dehydratase, wherein the peptide of interest is positioned C-terminally of both SEQ ID NO;
46 and the lantibiotic leader peptide. - View Dependent Claims (16)
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17. A nucleic acid molecule encoding a synthetic polypeptide, the nucleic acid molecule comprising:
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a first nucleotide sequence encoding a peptide selected from the group consisting of; an export signal peptide of a secretory (Sec) system, an export signal peptide of a Twin-Arginine Translocation (TAT) system, and an export signal peptide of a non-lantibiotic ATP Binding Cassette (ABC) export system; a second nucleotide sequence encoding a peptide selected from the group consisting of SEQ ID NO;
53 and SEQ ID NO;
54; anda third nucleotide sequence encoding a peptide of interest comprising a serine or threonine residue that can be posttranslationally dehydrated by dehydratase, wherein the third nucleotide sequence is positioned 3′
of both the first and second nucleotide sequence in the nucleic acid construct. - View Dependent Claims (18)
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19. A nucleic acid encoding a polypeptide that is synthetic, the polypeptide comprising:
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a first, export signal peptide that is SEQ ID NO;
46 or an export signal peptide for the secretory (Sec) system from Lactococcus lactis;
a second, leader peptide selected from the group consisting of SEQ ID NO;
53 and SEQ ID NO;
54; anda third, peptide of interest comprising at least one serine or threonine residue that can be posttranslationally dehydrated by dehydratase, wherein the peptide of interest is positioned C-terminally of both the export signal peptide and the leader peptide. - View Dependent Claims (20, 21, 22)
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Specification