Altered OspA of Borrelia burgdorferi

Abstract

Provided herein are OspA polypeptides from Lyme Disease-causing Borrelia having certain alteration(s). In one embodiment, the alteration(s) increase the conformational stability of the OspA polypeptide containing the alteration(s) while maintaining at least some of the antigenicity of the corresponding unaltered OspA polypeptide. In another embodiment, the altered OspA polypeptide has reduced cross-reactivity to hLFA-1, as compared to the corresponding unaltered OspA polypeptide.

What is claimed is:  
     
         1 . A polypeptide, comprising an amino acid sequence of OspA protein from  Borrelia burgdorferi  from about residue 139 to about residue 273 of  Borrelia burgdorferi  OspA protein, wherein the sequence includes at least one alteration selected from the group consisting of: residue 139 being methionine, residue 160 being tyrosine, residue 189 being methionine and combinations thereof, wherein the numbering corresponds to the numbering of SEQ ID NO: 7.  
     
     
         2 . The polypeptide of  claim 1 , wherein the polypeptide has increased conformational stability compared to the corresponding unaltered OspA polypeptide.  
     
     
         3 . The polypeptide of  claim 1 , wherein residue 160 is tyrosine and residue 189 is methionine.  
     
     
         4 . The polypeptide of  claim 1 , wherein residue 139 is methionine, residue 160 is tyrosine and residue 189 is methionine.  
     
     
         5 . The polypeptide of  claim 1 , wherein the polypeptide comprises residues 131 through 273 of the  Borrelia burgdorferi  OspA protein.  
     
     
         6 . The polypeptide of  claim 1 , wherein the polypeptide comprises residues 17 through 273 of the  Borrelia burgdorferi  OspA protein.  
     
     
         7 . The polypeptide of  claim 1 , wherein the polypeptide is derived from an OspA protein of a sensu stricto strain of  Borrelia burgdorferi.    
     
     
         8 . A polypeptide, comprising an amino acid sequence of OspA protein from a sensu stricto strain of  Borrelia burgdorferi  from about residue 160 to about residue 170, wherein the sequence includes at least one alteration selected from the group consisting of: residue 165 being phenylalanine, residue 166 being threonine, residue 170 being lysine and combinations thereof, wherein the numbering corresponds to the numbering of SEQ ID NO: 7.  
     
     
         9 . The polypeptide of  claim 8 , wherein cross-reactivity to an hLFA-1 molecule is reduced compared to the corresponding unaltered OspA polypeptide.  
     
     
         10 . The polypeptide of  claim 8 , wherein residue 165 is phenylalanine and residue 166 is threonine.  
     
     
         11 . The polypeptide of  claim 8 , wherein residue 165 is phenylalanine, residue 166 is threonine and residue 170 is lysine.  
     
     
         12 . The polypeptide of  claim 8 , wherein residue 139 is methionine, residue 160 is tyrosine and residue 189 is methionine.  
     
     
         13 . The polypeptide of  claim 8 , wherein the polypeptide comprises residues 150 through 180 of the  Borrelia burgdorferi  OspA protein.  
     
     
         14 . The polypeptide of  claim 8 , wherein the polypeptide comprises residues 17 through 273 of the  Borrelia burgdorferi  OspA protein.  
     
     
         15 . A polypeptide selected from the group consisting of: SEQ ID NO: 96, 98, 100, 102, 104, 106, 108, 110, 112, 114 and 116.  
     
     
         16 . A polynucleotide encoding an amino acid sequence of an OspA protein from  Borrelia burgdorferi  from about residue 139 to about residue 273, wherein the sequence encodes at least one alteration selected from the group consisting of: codon 139 encoding methionine, codon 160 encoding tyrosine, codon 189 encoding methionine and combinations thereof, wherein the numbering corresponds to the numbering of SEQ ID NO: 7.  
     
     
         17 . The polynucleotide of  claim 16 , wherein codon 160 encodes tyrosine and codon 189 encodes methionine.  
     
     
         18 . The polynucleotide of  claim 16 , wherein codon 139 encodes methionine, codon 160 encodes tyrosine and codon 189 encodes methionine.  
     
     
         19 . The polynucleotide of  claim 16 , wherein the encoded polypeptide comprises residues 131 through 273 of the  Borrelia burgdorferi  OspA protein.  
     
     
         20 . The polynucleotide of  claim 16 , wherein the encoded polypeptide comprises residues 17 through 273 of the  Borrelia burgdorferi  OspA protein.  
     
     
         21 . The polynucleotide of  claim 16 , wherein the encoded polypeptide corresponds to OspA protein of a sensu stricto strain of  Borrelia burgdorferi.    
     
     
         22 . A polynucleotide encoding an amino acid sequence of an OspA protein from  Borrelia burgdorferi  from about residue 160 to about residue 170, wherein the sequence encodes at least one alteration selected from the group consisting of: codon 165 encoding phenylalanine, codon 166 encoding threonine, codon 170 encoding lysine and combinations thereof, wherein the numbering corresponds to the numbering of SEQ ID NO: 7.  
     
     
         23 . The polynucleotide of  claim 22 , wherein codon 165 encodes phenylalanine and codon 166 encodes threonine.  
     
     
         24 . The polynucleotide of  claim 22 , wherein codon 165 encodes phenylalanine, codon 166 encodes threonine and codon 170 encodes lysine.  
     
     
         25 . A polynucleotide selected from the group consisting of: SEQ ID NO: 95, 97, 99, 101, 103, 105, 107, 109, 111, 113 and 115.  
     
     
         26 . A method of generating an altered  Borrelia burgdorferi  OspA polypeptide with increased conformational stability compared to the corresponding unaltered  Borrelia burgdorferi  OspA polypeptide comprising; 
 a) selecting a polynucleotide encoding a  Borrelia burgdorferi  OspA polypeptide that includes residues 139, 160 and 189, wherein the numbering corresponds to the numbering of SEQ ID NO: 7;    b) altering the polynucleotide such that residue 139 is methionine, residue 160 is tyrosine and residue 189 is methionine; and    c) expressing said altered polynucleotide;    thereby generating an altered  Borrelia burgdorferi  OspA polypeptide with increased conformational stability compared to the corresponding unaltered  Borrelia burgdorferi  OspA polypeptide.    
     
     
         27 . A method of generating an altered  Borrelia burgdorferi  OspA polypeptide with reduced cross-reactivity with an hLFA-1 molecule, compared to the corresponding unaltered  Borrelia burgdorferi  OspA polypeptide, comprising; 
 a) selecting a polynucleotide encoding an OspA polypeptide from  Borrelia burgdorferi  that includes residues 165, 166 and 170, wherein the numbering corresponds to the numbering of SEQ ID NO: 7;    b) altering the polynucleotide such that residue 165 is phenylalanine, residue 166 is threonine and residue 170 is lysine; and    c) expressing said altered polynucleotide;    thereby generating an altered  Borrelia burgdorferi  OspA polypeptide with reduced cross-reactivity with the hLFA-1 molecule compared to the corresponding unaltered  Borrelia burgdorferi  OspA polypeptide.    
     
     
         28 . Use of an altered OspA polypeptide for the manufacture of a medicament for immunizing a mammal, wherein the altered OspA polypeptide comprises: 
 a) an amino acid sequence of OspA protein from  Borrelia burgdorferi  from about residue 139 to about residue 273 of  Borrelia burgdorferi  OspA protein, wherein the sequence includes at least one alteration selected from the group consisting of: residue 139 being methionine, residue 160 being tyrosine, residue 189 being methionine and combinations thereof, wherein the polypeptide has increased conformational stability compared to a corresponding wild type OspA polypeptide; and/or    b) an amino acid sequence of OspA protein from  Borrelia burgdorferi  from about residue 160 to about residue 170, wherein the sequence includes at least one alteration selected from the group consisting of: residue 165 being phenylalanine, residue 166 being threonine, residue 170 being lysine and combinations thereof, wherein the polypeptide has decreased cross-reactivity to hLFA-1 compared to a corresponding wild type OspA polypeptide;    wherein the numbering corresponds to the numbering of SEQ ID NO: 7.    
     
     
         29 . A polypeptide, comprising an amino acid sequence of OspA protein from  Borrelia burgdorferi  from about residue 160 to about residue 170, wherein the sequence includes at least two alterations selected from the group consisting of: residue 165 being phenylalanine, residue 166 being threonine and residue 170 being lysine, wherein the numbering corresponds to the numbering of SEQ ID NO: 7.  
     
     
         30 . A chimeric polypeptide, comprising: 
 a) an amino acid sequence of a first OspA polypeptide from about residue 1 to about residue 164 from a first strain of  Borrelia burgdorferi;      b) an amino acid sequence of a second OspA polypeptide from about residue 165 to about residue 179 from a second strain of  Borrelia burgdorferi , wherein said second strain is a different strain from said first strain;    c) an amino acid sequence of a third OspA polypeptide from about residue 180 to about residue 216 from a third strain of  Borrelia burgdorferi , wherein said third strain is a different strain from said second strain;    d) an amino acid sequence of a fourth OspA polypeptide from about residue 217 to about residue 273 from a fourth strain of  Borrelia burgdorferi , wherein said fourth strain is a different strain from said third strain;    wherein the sequence includes at least one alteration selected from the group consisting of: residue 139 being methionine, residue 160 being tyrosine, residue 189 being methionine and combinations thereof, wherein the numbering corresponds to the numbering of SEQ ID NO: 7.