Follistatin-related protein zfsta2

Abstract

The present invention relates to polynucleotide and polypeptide molecules for zfsta2, a novel member of the follistatin family. The polypeptides, and polynucleotides encoding them are useful for binding to members of the TGF-β family and mediating central nervous system, reproductive, hematopoietic and bone-related activities. The present invention also includes antibodies to the zfsta2 polypeptides.

What is claimed is:  
     
         1 . An isolated polypeptide comprising a follistatin homology domain, wherein said follistatin homology domain comprises amino acid residues 65 to 133 of the amino acid sequence of SEQ ID NO:2.  
     
     
         2 . An isolated polypeptide of  claim 1 , wherein said polypeptide further comprises an alpha helical linker region that resides in a carboxyl-terminal position relative to said follistatin homology domain, wherein said alpha helical linker region comprises amino acid residues 134 to 174 of the amino acid sequence of SEQ ID NO:2.  
     
     
         3 . An isolated polypeptide of  claim 2 , wherein said polypeptide further comprises a calmodulin homology domain that resides in a carboxyl-terminal position relative to said alpha helical linker region, wherein said calmodulin homology domain comprises amino acid residues 175 to 250 of the amino acid sequence of SEQ ID NO:2.  
     
     
         4 . An isolated polypeptide of  claim 2 , wherein said polypeptide further comprises two I-set Ig domains that reside in a carboxyl-terminal position relative to said calmodulin homology domain, wherein said I-set Ig domains comprise amino acid residues 251 to 431 of the amino acid sequence of SEQ ID NO:2.  
     
     
         5 . An isolated polypeptide of  claim 2 , wherein said polypeptide further comprises a carboxy-terminal domain that resides in a carboxyl-terminal position relative to said I-set Ig domains, wherein said carboxy-terminal domain comprises amino acid residues 433 to 847 of said amino acid sequence of SEQ ID NO:2.  
     
     
         6 . An isolated polypeptide of  claim 1 , wherein said polypeptide further comprises a hydrophilic linker region that resides in an amino-terminal position relative to said follistatin homology domain, wherein said hydrophobic linker region comprises amino acid residues 21 to 64 of the amino acid sequence of SEQ ID NO:2.  
     
     
         7 . An isolated polypeptide of  claim 6 , wherein said polypeptide further comprises a secretory signal sequence that resides in an amino-terminal position relative to said hydrophobic linker region, wherein said secretory signal sequence comprises amino acid residues 1 to 20 of the amino acid sequence of SEQ ID NO:2.  
     
     
         8 . An isolated polypeptide having an amino acid sequence that is at least 70% identical to the amino acid sequence of SEQ ID NO:2, wherein said isolated polypeptide specifically binds with an antibody to which a polypeptide having the amino acid sequence of SEQ ID NO:2 specifically binds.  
     
     
         9 . An isolated polypeptide of  claim 8 , wherein said isolated polypeptide has an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:2.  
     
     
         10 . An isolated polypeptide of  claim 8 , wherein said isolated polypeptide has an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2.  
     
     
         11 . An isolated polypeptide of  claim 8 , wherein any difference between said amino acid sequence and said corresponding amino acid sequence of SEQ ID NO:2 is due to one or more conservative amino acid substitutions.  
     
     
         12 . An isolated polypeptide of  claim 8 , wherein the amino acid percent identity is determined using a FASTA program with ktup=1, gap opening penalty=10, gap extension penalty=1, and substitution matrix=blosum62, with other parameters set as default.  
     
     
         13 . An isolated polypeptide comprising the amino acid sequence of SEQ ID NO:2.  
     
     
         14 . An isolated polypeptide selected from the group consisting of: 
 a) a polypeptide consisting of the sequence of amino acid residues from residue 21 to residue 64 of SEQ ID NO:2;    b) a polypeptide consisting of the sequence of amino acid residues from residue 65 to residue 133 of SEQ ID NO :2;    c) a polypeptide consisting of the sequence of amino acid residues from residue 134 to residue 174 of SEQ ID NO:2;    d) a polypeptide consisting of the sequence of amino acid residues from residue 175 to residue 250 of SEQ ID NO:2;    e) a polypeptide consisting of the sequence of amino acid residues from residue 251 to residue 334 of SEQ ID NO:2;    f) a polypeptide consisting of the sequence of amino acid residues from residue 335 to residue 432 of SEQ ID NO:2;    g) a polypeptide consisting of the sequence of amino acid residues from residue 433 to residue 847 of SEQ ID NO:2;    h) a polypeptide consisting of the sequence of amino acid residues from residue 251 to residue 432 of SEQ ID NO:2;    i) a polypeptide consisting of the sequence of amino acid residues from residue 65 to residue 174 of SEQ ID NO:2;    j) a polypeptide consisting of the sequence of amino acid residues from residue 65 to residue 250 of SEQ ID NO: 2;    k) a polypeptide consisting of the sequence of amino acid residues from residue 65 to residue 334 of SEQ ID NO:2;    l) a polypeptide consisting of the sequence of amino acid residues from residue 65 to residue 847 of SEQ ID NO:2;    m) a polypeptide consisting of the sequence of amino acid residues from residue 134 to residue 250 of SEQ ID NO: 2;    n) a polypeptide consisting of the sequence of amino acid residues from residue 134 to residue 334 of SEQ ID NO:2;    o) a polypeptide consisting of the sequence of amino acid residues from residue 134 to residue 432 of SEQ ID NO:2;    p) a polypeptide consisting of the sequence of amino acid residues from residue 134 to residue 847 of SEQ ID NO:2;    q) a polypeptide consisting of the sequence of amino acid residues from residue 175 to residue 334 of SEQ ID NO: 2;    r) a polypeptide consisting of the sequence of amino acid residues from residue 175 to residue 432 of SEQ ID NO:2; and    s) a polypeptide consisting of the sequence of amino acid residues from residue 175 to residue 847 of SEQ ID NO: 2.    
     
     
         15 . An isolated polypeptide according to  claim 1 , further comprising an affinity tag or binding domain.  
     
     
         16 . A fusion protein comprising a secretory signal sequence having the amino acid sequence of amino acid residues 1-20 of SEQ ID NO:2, wherein said secretary signal sequence is operably linked to an additional polypeptide.  
     
     
         17 . A fusion protein consisting essentially of a first portion and a second portion joined by a peptide bond, said first portion comprising a polypeptide according to  claim 1;  and 
 said second portion comprising another polypeptide.  
 
     
     
         18 . An isolated polynucleotide molecule that encodes a polypeptide according to  claim 1 .  
     
     
         19 . An isolated polynucleotide molecule according to  claim 18 , encoding a polypeptide further comprising an alpha helical linker region that resides in a carboxyl-terminal position relative to said follistatin homology domain, wherein said alpha helical linker region comprises amino acid residues 134 to 174 of the amino acid sequence of SEQ ID NO:2.  
     
     
         20 . An isolated polynucleotide of  claim 19 , wherein said polynucleotide encodes a polypeptide further comprising a calmodulin homology domain that resides in a carboxyl-terminal position relative to said alpha helical linker region, wherein said calmodulin homology domain comprises amino acid residues 175 to 250 of the amino acid sequence of SEQ ID NO:2.  
     
     
         21 . An isolated polynucleotide of  claim 20 , wherein said polynucleotide encodes a polypeptide further comprising two I-set Ig domains that reside in a carboxyl-terminal position relative to said calmodulin homology domain, wherein said I-set Ig domains comprise amino acid residues 251 to 431 of the amino acid sequence of SEQ ID NO:2.  
     
     
         22 . An isolated polynucleotide of  claim 21 , wherein said polynucleotide encodes a polypeptide further comprising a carboxy-terminal domain that resides in a carboxyl-terminal position relative to said I-set Ig domains, wherein said carboxy-terminal domain comprises amino acid residues 433 to 847 of said amino acid sequence of SEQ ID NO:2.  
     
     
         23 . An isolated polynucleotide according to  claim 22 , wherein said polypeptide further comprises an affinity tag or binding domain.  
     
     
         24 . An isolated polynucleotide molecule, wherein said polynucleotide molecule is a degenerate nucleotide sequence encoding a polypeptide according to  claim 1 .  
     
     
         25 . An isolated polynucleotide encoding a polypeptide having an amino acid sequence that is at least 70% identical to the amino acid sequence of SEQ ID NO:2, wherein said isolated polypeptide specifically binds with an antibody to which a polypeptide having the amino acid sequence of SEQ ID NO:2 specifically binds.  
     
     
         26 . An isolated polynucteotide of  claim 25 , wherein said isolated polypeptide has an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:2.  
     
     
         27 . An isolated polynucleotide of  claim 26 , wherein said isolated polypeptide has an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2.  
     
     
         28 . An isolated polynucleotide of  claim 27 , wherein any difference between said amino acid sequence and said corresponding amino acid sequence of SEQ ID NO:2 is due to one or more conservative amino acid substitutions.  
     
     
         29 . An isolated polynucleotide of  claim 27 , wherein the amino acid percent identity is determined using a FASTA program with ktup=1, gap opening penalty=10, gap extension penalty=1, and substitution matrix=blosum62, with other parameters set as default.  
     
     
         30 . An isolated polynucleotide molecule comprising the nucleotide sequence of nucleotides 58 to 3006 of SEQ ID NO:1.  
     
     
         31 . An isolated polynucleotide molecule of SEQ ID NO:1.  
     
     
         32 . An isolated polynucleotide selected from the group consisting of: 
 a) a polynucleotide consisting of nucleotides 58-117 of SEQ ID NO:1;    b) a polynucleotide consisting of nucleotides 118-249 of SEQ ID NO:1;    c) a polynucleotide consisting of nucleotides 250-456 of SEQ ID NO:1;    d) a polynucleotide consisting of nucleotides 457-579 of SEQ ID NO:1;    e) a polynucleotiqe consisting of nucleotides 580-810 of SEQ ID NO:1;    f) a polynucleotide consisting of nucleotides 811-1059 of SEQ ID NO:1;    g) a polynucleotide consisting of nucleotides 1060-1353 of SEQ ID NO:1;    h) a polynucleotide consisting of nucleotides 1354-3006 of SEQ ID NO:1;    i) a polynucleotide consisting of nucleotides 250-579 of SEQ ID NO:1;    j) a polynucleotide consisting of nucleotides 250-810 of SEQ ID NO:1;    k) a polynucleotide consisting of nucleotides 250-1059 of SEQ ID NO:1;    l) a polynucleotide consisting of nucleotides 250-1353 of SEQ ID NO:1;    m) a polynucleotide consisting of nucleotides 250-3006 of SEQ ID NO:1;    n) a polynucleotide consisting of nucleotides 457-810 of SEQ ID NO:1;    o) a polynucleotide consisting of nucleotides 457-1059 of SEQ ID NO:1;    p) a polynucleotide consisting of nucleotides 457-1353 of SEQ ID NO:1;    q) a polynucleotide consisting of nucleotides 457-3006 of SEQ ID NO:1;    r) a polynucleotide consisting of nucleotides 580-1059 of SEQ ID NO:1;    s) a polynucleotide consisting of nucleotides 580-1353 of SEQ ID NO:1;    t) a polynucleotide consisting of nucleotides 580-3006 of SEQ ID NO:1; and    u) a polynucleotide consisting of nucleotides 811-1353 of SEQ ID NO:1.    
     
     
         33 . A polynucleotide encoding a fusion protein comprising a secretory signal sequence having the amino acid sequence of amino acid residues 1-20 of SEQ ID NO:2, wherein said secretory signal sequence is operably linked to an additional polypeptide.  
     
     
         34 . A polynucleotide molecule encoding a fusion protein consisting essentially of a first portion and a second portion joined by a peptide bond, said first portion comprising a polypeptide according to  claim 1;  and 
 said second portion comprising another polypeptide.  
 
     
     
         35 . An expression vector comprising the following operably linked elements: 
 a transcription promoter;    a polynucleotide molecule that encodes a polypeptide according to  claim 1;  and    a transcription terminator.    
     
     
         36 . An expression vector according to  claim 35  further comprising a secretory signal sequence operably linked to said DNA segment.  
     
     
         37 . An expression vector according to  claim 35 , wherein said polynucleotide encodes a polypeptide covalently linked amino terminally or carboxy terminally to an affinity tag.  
     
     
         38 . A cultured cell into which has been introduced an expression vector comprising the following operably linked elements: 
 a transcription promoter;    a polynucleotide molecule that encodes a polypeptide according to  claim 1;  and    a transcription terminator, wherein said cultured cell expresses said polypeptide encoded by said polynucleotide segment.    
     
     
         39 . A method of producing a polypeptide comprising: 
 culturing a cell into which has been introduced an expression vector comprising the following operably linked elements:    a transcription promoter;    a polynucleotide molecule that encodes a polypeptide according to  claim 1;  and    a transcription terminator;    whereby said cell expresses said polypeptide encoded by said polynucleotide segment; and    recovering said expressed polypeptide.    
     
     
         40 . An antibody or antibody fragment that specifically binds to a polypeptide according to  claim 1 .  
     
     
         41 . An antibody according to  claim 40 , wherein said antibody is selected from the group consisting of: 
 a) polyclonal antibody;    b) murine monoclonal antibody;    c) humanized antibody derived from b); and    d) human monoclonal antibody.    
     
     
         42 . An antibody fragment according to  claim 41 , wherein said antibody fragment is selected from the group consisting of F(ab′), F(ab), Fab′, Fab, Fv, scFv, and minimal recognition unit.  
     
     
         43 . An anti-idiotype antibody that specifically binds to said antibody of  claim 40   
     
     
         44 . A polypeptide according to  claim 1 , in combination with a pharmaceutically acceptable vehicle.