Secreted salivary zsig63 polypeptide

Abstract

The present invention relates to polynucleotide and polypeptide molecules for zsig63, a novel secreted salivary protein. The polypeptides, and polynucleotides encoding them, may exhibit anti-microbial activity and may be used in the study or treatment of microbial infections. The polynucleotides encoding zsig63, are located on chromosome 4, and can be used to identify a region of the genome associated with human disease states. The present invention also includes antibodies to the zsig63 polypeptides.

1 . An isolated polynucleotide encoding a zsig63 polypeptide comprising a sequence of amino acid residues that is at least 90% identical to an amino acid sequence selected from the group consisting of: 
 (a) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 16 (Arg) to amino acid number 37 (Ser);    (b) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 38 (Leu) to amino acid number 126 (Ala);    (c) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 127 (Pro) to amino acid number 219 (Gln);    (d) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 16 (Arg) to amino acid number 219 (Gln); and    (e) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 1 (Met) to amino acid number 219 (Gln).    
     
     
         2 . An isolated polynucleotide according to  claim 1 , wherein the zsig63 polypeptide comprises a sequence of amino acid residues selected from the group consisting of: 
 (a) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 16 (Arg) to amino acid number 37 (Ser);    (b) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 38 (Leu) to amino acid number 126 (Ala);    (c) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 127 (Pro) to amino acid number 219 (Gln);    (d) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 16 (Arg) to amino acid number 219 (Gln); and    (e) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 1 (Met) to amino acid number 219 (Gln).    
     
     
         3 . An isolated polynucleotide according to  claim 1 , wherein the polynucleotide is selected from the group consisting of: 
 (a) a polynucleotide sequence as shown in SEQ ID NO: 1 from nucleotide 173 to nucleotide 784;    (b) a polynucleotide sequence as shown in SEQ ID NO: 1 from nucleotide 128 to nucleotide 784; and    (c) a polynucleotide sequence complementary to (a) or (b).    
     
     
         4 . An isolated polynucleotide sequence according to  claim 1 , wherein the polynucleotide comprises nucleotide 1 to nucleotide 657 of SEQ ID NO:3.  
     
     
         5 . An expression vector comprising the following operably linked elements: 
 a transcription promoter;    a DNA segment encoding a zsig63 polypeptide comprising an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:2 from amino acid number 16 (Arg) to amino acid number 219 (Gln); and    a transcription terminator.    
     
     
         6 . An expression vector according to  claim 5 , further comprising a secretory signal sequence operably linked to the DNA segment.  
     
     
         7 . A cultured cell into which has been introduced an expression vector according to  claim 5 , wherein the cell expresses a polypeptide encoded by the DNA segment.  
     
     
         8 . A DNA construct encoding a fusion protein, the DNA construct comprising: 
 a first DNA segment encoding a polypeptide selected from the group consisting of:    (a) the amino acid sequence of SEQ ID NO: 2 from residue number 1 (Met) to residue number 15 (Ala);    (b) the amino acid sequence of SEQ ID NO: 2 from residue number 16 (Arg) to residue number 37 (Ser);    (c) the amino acid sequence of SEQ ID NO: 2 from residue number 38 (Leu) to residue number 126 (Ala);    (d) the amino acid sequence of SEQ ID NO: 2 from residue number 127 (Pro) to residue number 219 (Gln); and    (e) the amino acid sequence of SEQ ID NO:2 from residue number 16 (Arg) to residue number 219 (Gin); and    at least one other DNA segment encoding an additional polypeptide,    wherein the first and other DNA segments are connected in-frame; and    encode the fusion protein.    
     
     
         9 . A fusion protein produced by a method comprising: 
 culturing a host cell into which has been introduced a vector comprising the following operably linked elements:    (a) a transcriptional promoter;    (b) a DNA construct encoding a fusion protein according to  claim 8;  and    (c) a transcriptional terminator; and    recovering the protein encoded by the DNA segment.    
     
     
         10 . An isolated zsig63 polypeptide comprising a sequence of amino acid residues that is at least 90% identical to an amino acid sequence selected from the group consisting of: 
 (a) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 16 (Arg) to amino acid number 37 (Ser);    (b) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 38 (Leu) to amino acid number 126 (Ala);    (c) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 127 (Pro) to amino acid number 219 (Gln);    (d) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 16 (Arg) to amino acid number 219 (Gln); and    (e) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 1 (Met) to amino acid number 219 (Gln).    
     
     
         11 . An isolated zsig63 polypeptide according to  claim 10 , wherein the polypeptide comprises a sequence of amino acid residues selected from the group consisting of: 
 (a) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 16 (Arg) to amino acid number 37 (Ser);    (b) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 38 (Leu) to amino acid number 126 (Ala);    (c) the amino acid sequence as shown in SEQ ID NO: 2 from amino acid number 127 (Pro) to amino acid number 219 (Gln);    (d) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 16 (Arg) to amino acid number 219 (Gln); and    (e) the amino acid sequence as shown in SEQ ID NO:2 from amino acid number 1 (Met) to amino acid number 219 (Gln).    
     
     
         12 . A method of producing a zsig63 polypeptide comprising: 
 culturing a cell according to  claim 7;  and    isolating the zsig63 polypeptide produced by the cell.    
     
     
         13 . A method of detecting, in a test sample, the presence of an antagonist of zsig63 protein activity, comprising: 
 transfecting a zsig63-responsive cell, with a reporter gene construct that is responsive to a zsig63-stimulated cellular pathway; and    producing a zsig63 polypeptide by the method of  claim 12;  and    adding the zsig63 polypeptide to the cell, in the presence and absence of a test sample; and    comparing levels of response to the zsig63 polypeptide, in the presence and absence of the test sample, by a biological or biochemical assay; and    determining from the comparison, the presence of the antagonist of zsig63 activity in the test sample.    
     
     
         14 . A method of detecting, in a test sample, the presence of an agonist of zsig63 protein activity, comprising: 
 transfecting a zsig63-responsive cell, with a reporter gene construct that is responsive to a zsig63-stimulated cellular pathway; and    adding a test sample; and    comparing levels of response in the presence and absence of the test sample, by a biological or biochemical assay; and    determining from the comparison, the presence of the agonist of zsig63 activity in the test sample.    
     
     
         15 . A method of producing an antibody to zsig63 polypeptide comprising the following steps in order: 
 inoculating an animal with a polypeptide selected from the group consisting of:    (a) a polypeptide consisting of 9 to 204 amino acids, wherein the polypeptide consists of a contiguous sequence of amino acids in SEQ ID NO:2 from amino acid number 16 (Ala) to amino acid number 219 (Gln); and    (b) a polypeptide according to  claim 10;     (c) a polypeptide comprising amino acid number 16 (Arg) to 37 (Ser) of SEQ ID NO:2;    (d) a polypeptide comprising amino acid number 38 (Leu) to 126 (Ala) of SEQ ID NO:2;    (e) a polypeptide comprising amino acid number 127 (Pro) to 219 (Gln) of SEQ ID NO:2;    (f) a polypeptide comprising amino acid number 16 (Arg) to amino acid number 219 (Gln) of SEQ ID NO:2;    (g) a polypeptide comprising amino acid number 1 (Met) to amino acid number 219 (Gln) of SEQ ID NO:2;    (h) a polypeptide comprising amino acid number 14 (Phe) to 19 (Arg) of SEQ ID NO:2;    (i) a polypeptide comprising amino acid number 16 (Arg) to 21 (Phe) of SEQ ID NO:2;    (j) a polypeptide comprising amino acid number 24 (Gly) to 29 (Asp) of SEQ ID NO:2;    (k) a polypeptide comprising amino acid number 25 (Glu) to 30 (Asp) of SEQ ID NO:2;    (l) a polypeptide comprising amino acid number 187Glu) to 192 (Glu) of SEQ ID NO:2;    (m) a polypeptide comprising amino acid number 24 (Gly) to 33 (Pro) of SEQ ID NO:2;    (n) a polypeptide comprising amino acid number 17 (Lys) to 33 (Pro) of SEQ ID NO:2;    (o) a polypeptide comprising amino acid number 66 (Thr) to 73 (Pro) of SEQ ID NO:2;    (p) a polypeptide comprising amino acid number 103 (Pro) to 108 (Gly) of SEQ ID NO:2;    (q) a polypeptide comprising amino acid number 190 (Ala) to 197 (Glu) of SEQ ID NO:2;    (r) a polypeptide comprising amino acid number 202 (Lys) to 215 (Gly) of SEQ ID NO:2; and    (s) a polypeptide comprising amino acid number 190 (Ala) to 215 (Glu) of SEQ ID NO:2; and    wherein the polypeptide elicits an immune response in the animal to produce the antibody; and    isolating the antibody from the animal.    
     
     
         16 . An antibody produced by the method of  claim 15 , which binds to a zsig63 polypeptide.  
     
     
         17 . The antibody of  claim 16 , wherein the antibody is a monoclonal antibody.  
     
     
         18 . An antibody that binds to a polypeptide of  claim 10.