Chimeric autoprocessing polypeptides and uses thereof

Abstract

A chimeric polypeptide comprising an autoprocessing segment having an amino acid sequence being capable of auto-cleavage, a polynucleotide encoding such a polypeptide, and uses of such a polypeptide and such a polynucleotide are provided. a polynucleotide are provided.

1 . A chimeric polypeptide comprising an autoproces sing segment having an amino acid sequence set forth by SEQ ID NO: 37. 
     
     
         2 . The chimeric polypeptide of  claim 1 , wherein said autoprocessing segment is derived from a protein of an organism belonging to a genus selected from the group consisting of  Leptospira interrogans.    
     
     
         3 . The chimeric polypeptide of  claim 1 , wherein said auto-cleavage results in removal of a segment of the chimeric polypeptide adjacent to an amino terminal end or a carboxy terminal end of said autoprocessing segment. 
     
     
         4 . The chimeric polypeptide of  claim 1 , wherein said auto-cleavage results in auto-splicing. 
     
     
         5 . The chimeric polypeptide of  claim 4 , wherein said auto-splicing is auto-splicing of segments of the chimeric polypeptide flanking said autoprocessing segment. 
     
     
         6 . The chimeric polypeptide of  claim 1 , further comprising an affinity tag capable of specifically binding a substrate. 
     
     
         7 . The chimeric polypeptide of  claim 6 , wherein said affinity tag is a maltose-binding domain or a chitin-binding domain. 
     
     
         8 . The chimeric polypeptide of  claim 6 , wherein said substrate is selected from the group consisting of a molecule, a compound, a virus, and a cell. 
     
     
         9 . A polynucleotide encoding a chimeric polypeptide comprising an autoprocessing segment having an amino acid sequence set forth by SEQ ID NO: 37, the polypeptide being capable of auto-cleavage. 
     
     
         10 . The polynucleotide of  claim 9 , further comprising a promoter sequence being for directing expression of the chimeric polypeptide in an expression system. 
     
     
         11 . The polynucleotide of  claim 10 , wherein said promoter sequence is inducible by isopropyl beta-D-thiogalactoside. 
     
     
         12 . A nucleic acid construct comprising the polynucleotide of  claim 10 . 
     
     
         13 . A method of purifying a protein, the method comprising:
 (a) generating a chimeric polypeptide including an autoproces sing segment having an amino acid sequence set forth by SEQ ID NO: 37, said autoprocessing segment being terminally attached to, or flanked by, an amino acid sequence of the protein, said chimeric polypeptide being capable of auto-cleavage when subjected to suitable conditions to thereby remove said amino acid sequence of the protein from said chimeric polypeptide thereby generating the protein;   (b) immobilizing said chimeric polypeptide to a support; and   (c) subjecting said chimeric polypeptide to said suitable conditions, thereby purifying the protein.   
     
     
         14 . The method of  claim 13 , further comprising the step of separating the protein from said autoprocessing segment following step (c). 
     
     
         15 . The method of  claim 13 , wherein said support includes an antibody or antibody fragment capable of specifically binding said autoprocessing segment, and whereas said immobilizing is via said autoprocessing segment. 
     
     
         16 . The method of  claim 13 , wherein said chimeric polypeptide further includes an affinity tag sequence, and whereas said immobilizing is via said affinity tag sequence. 
     
     
         17 . The method of  claim 16 , wherein said support includes a specific ligand of said affinity tag sequence, and whereas said immobilizing is via said specific ligand of said affinity tag sequence.