Moraxella catarrhalis Proteins

Abstract

Novel antigens of B. catarrhalis are provided, together with their use in vaccines as well as methods of diagnosing and/or detection.

1 . A purified protein that is a  Branhamella  ( Moraxella )  catarrhalis  antigen, wherein said protein has an apparent molecular mass of about 30 kDa as determined by SDS-PAGE, and the following N-terminal sequence: 
       
         
           
                 
                 
                 
               
                   NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV. 
                   (SEQ ID NO: 2) 
                     
                 
             
                
               
            
           
         
       
     
     
         2 . A protein preparation comprising an outer membrane  Branhamella  ( Moraxella )  catarrhalis  protein that has an apparent molecular mass of about 30 kDa, as determined by SDS-PAGE, and has the N-terminal sequence: NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV (SEQ ID NO: 2), wherein when said preparation is subjected to analytical SDS-PAGE, followed by staining of the SDS-PAGE gel, said about 30 kDa protein is the most abundant  Branhamella  ( Moraxella )  catarrhalis  protein in said preparation and wherein mice immunized with said 30 kDa protein and challenged with said bacteria have an enhanced clearance of said bacteria from a bronchoalveolar lavage and their lungs as compared to that of non-immunized mice. 
     
     
         3 . The protein preparation of  claim 2 , wherein said protein that has an apparent molecular mass of about 30 kDa is producible by a process comprising:
 (a) growing a culture of  Branhamella  ( Moraxella )  catarrhalis  bacterial cells,   (b) washing said bacterial cells by resuspending said cells in phosphate buffered saline (PBS) and centrifuging to pellet said cells;   (c) extracting outer membrane proteins from the washed bacterial cells by the addition of a zwittergent and subsequent ethanol precipitation;   (d) purifying said about 30 kDa outer membrane protein by ion exchange chromatography of the extracted outer membrane proteins of part (c) on an ion exchange column that has an ion exchange like that of a Q2 column and pooling the fractions that contain said 30 kDa protein; and   (e) purifying said protein by preparative gel electrophoresis under conditions   that separate the proteins that are being eluted into at least two fractions based on a protein's molecular mass, and selecting the fraction(s) that contain said protein that has said apparent molecular mass of about 30 kDa.   
     
     
         4 . An immunogenic composition comprising a purified protein that is a  Branhamella  ( Moraxella )  catarrhalis  antigen, wherein said protein has an apparent molecular mass of about 30 kDa as determined by SDS-PAGE, and the following N-terminal sequence: 
       
         
           
                 
                 
                 
               
                   NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV. 
                   (SEQ ID NO: 2) 
                     
                 
             
                
               
            
           
         
       
     
     
         5 . The immunogenic composition of  claim 4 , wherein said composition further comprises at least one other  Branhamella  ( Moraxella )  catarrhalis  antigen. 
     
     
         6 . A vaccine comprising the immunogenic composition as claimed in  claim 4  or  claim 5 . 
     
     
         7 . A method for the prophylaxis or treatment of a respiratory infection, said method comprising vaccinating a subject with the vaccine of  claim 6 . 
     
     
         8 . The method of  claim 7 , wherein said respiratory infection is otitis media. 
     
     
         9 . An antibody raised against the protein of  claim 1 . 
     
     
         10 . A method of detecting or diagnosing  Branhamella  ( Moraxella )  catarrhalis,  wherein said method comprises:
 a) contacting one or more antibodies with a sample to be tested, wherein said antibodies are antibodies to a  Branhamella  ( Moraxella )  catarrhalis  protein antigen, wherein said protein antigen has an apparent molecular mass of about 30 kDa as determined by SDS-PAGE, and the following N-terminal sequence: NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV (SEQ ID NO:2); and   b) detecting or diagnosing the presence of said  Branhamella  ( Moraxella )  catarrhalis  by detecting the binding of said antibodies to said antigen.   
     
     
         11 . The method of  claim 10 , wherein said  Branhamella  ( Moraxella )  catarrhalis  is detected or diagnosed in vitro. 
     
     
         12 . A kit for use in detecting or diagnosing  Branhamella  ( Moraxella )  catarrhalis,  said kit comprising a purified protein that is a  Branhamella  ( Moraxella )  catarrhalis  antigen, wherein said protein has an apparent molecular mass of about 30 kDa as determined by SDS-PAGE, and the following N-terminal sequence: NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV (SEQ ID NO:2), or an antibody to said protein. 
     
     
         13 . A method of inducing an immune response against  Branhamella  ( Moraxella )  catarrhalis  in a subject, said method comprising immunizing said subject with the vaccine of  claim 6 . 
     
     
         14 . An isolated nucleic acid molecule comprising:
 (i) a DNA or RNA sequence coding for a protein that is a  Branhamella  ( Moraxella )  catarrhalis  antigen, wherein said protein has an apparent molecular mass of about 30 kDa as determined by SDS-PAGE, and the following N-terminal sequence: NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV (SEQ ID NO:2), or,   (ii) a DNA or RNA sequence coding for an antigenic fragment of said protein of part (i);   (iii) a DNA or RNA sequence complementary to any of the sequences of (i); or   (iv) a DNA or RNA sequence that has at least 75% identity with any of the coding sequence of (i) or the complementary sequence of (ii).   
     
     
         15 . The isolated nucleic acid molecule of  claim 14 , wherein said molecule comprises a DNA or RNA sequence that has at least 95% identity with any of the coding sequence of (i) or the complementary sequence of (ii). 
     
     
         16 . A vector comprising the nucleic acid molecule of  claim 14 . 
     
     
         17 . A host cell transformed or transfected with the vector of  claim 16 . 
     
     
         18 . A method for the production of a protein, said method comprising expressing a nucleic acid encoding said protein in a host cell that has been transformed or transfected with said nucleic acid encoding said protein, wherein said protein is a  Branhamella  ( Moraxella )  catarrhalis  antigen that has an apparent molecular mass of about 30 kDa, as determined by SDS-PAGE, and that has the following N-terminal sequence: 
       
         
           
                 
                 
                 
               
                   NVVTNTGATVVDGTRTIFSTLVKPAAVVAAV. 
                   (SEQ ID NO: 2)