Acetyl coa carboxylase 2 sequences and methods

Abstract

The present invention relates generally to novel nucleotide and amino acid sequences, and more particularly to novel human acetyl CoA carboxylase 2 (ACC2) and rat ACC2 sequences. The sequences provided herein can be expressed in a recombinant format. Methods of isolating the ACC2 sequence are also provided, which can be employed to isolate any ACC sequence. The ACC2 sequences can be employed in therapeutic applications to diagnose or treat a condition associated with ACC2. The invention also relates to the identification of modulators of ACC activity using the recombinant human ACC2 enzyme as the screening target.

1 - 7 . (canceled)  
     
     
         8 . An isolated polypeptide comprising an amino acid sequence selected from the group consisting of: 
 (a) a polypeptide comprising SEQ ID NO:13;    (b) a polypeptide comprising amino acids 2 to 2458 of SEQ ID NO:13, wherein amino acids 2 to 2458 comprise a polypeptide of SEQ ID NO:13 minus the start methionine; and    (c) a polypeptide comprising amino acids 1 to 2458 of SEQ ID NO:13.    
     
     
         9 . The isolated polypeptide of  claim 8 , wherein the polypeptide comprises two or more sequential amino acid deletions from one or both of: 
 (a) the COOH-terminus of the polypeptide; and    (b) the NH 2 -terminus of the polypeptide.    
     
     
         10 . A method of identifying a compound that modulates the activity of the polypeptide of  claim 8 , the method comprising: 
 (a) determining the activity of the polypeptide of  claim 8  in the absence of a test compound;    (b) determining the activity of the polypeptide in the presence of a test compound; and    (c) comparing the activity of the polypeptide in the presence of the test compound with the activity of the polypeptide in the absence of the test compound,    wherein a change in the activity of the polypeptide in the presence of the test compound relative to the activity of the polypeptide in the absence of the test compound indicates that the compound that modulates the activity of the polypeptide.    
     
     
         11 . An isolated antibody which specifically binds to the polypeptide of  claim 8 .  
     
     
         12 . The antibody of  claim 11 , wherein the antibody is selected from the group consisting of a chimeric antibody, a single chain antibody, a Fab fragment, and a humanized antibody.  
     
     
         13 . An isolated nucleic acid molecule comprising a polynucleotide having a nucleotide sequence selected from the group consisting of: 
 (a) a polynucleotide encoding an ACC2 polypeptide comprising SEQ ID NO:16;    (b) an isolated polynucleotide encoding a rat ACC2 polypeptide comprising amino acids 2 to 2455 of SEQ ID NO:16 minus the start methionine;    (c) an isolated polynucleotide encoding a rat ACC2 polypeptide comprising amino acids 1 to 2455 of SEQ ID NO:16 including the start codon;    (d) the cDNA of ATCC Deposit No. PTA-6054; and    (e) a polynucleotide capable of hybridizing under stringent conditions to the polynucleotide specified in (a)-(d), wherein the polynucleotide does not hybridize under stringent conditions to a nucleic acid molecule having a nucleotide sequence of only A residues or of only T residues.    
     
     
         14 . The isolated nucleic acid molecule of  claim 13 , wherein the polynucleotide comprises the nucleotide sequence of SEQ ID NO:15.  
     
     
         15 . A polynucleotide that is complementary to the isolated nucleic acid molecule of  claim 13 .  
     
     
         16 . A vector comprising the isolated nucleic acid molecule of  claim 13 .  
     
     
         17 . A host cell comprising the vector of  claim 16 .  
     
     
         18 . The host cell of  claim 17 , wherein the host cell is a mammalian host cell.  
     
     
         19 . A method of making an isolated polypeptide comprising: 
 (a) culturing the recombinant host cell of  claim 17  under conditions such that the polypeptide is expressed; and    (b) recovering the polypeptide.    
     
     
         20 . An isolated polypeptide comprising an amino acid sequence selected from the group consisting of: 
 (a) a polypeptide comprising SEQ ID NO:16;    (b) a polypeptide comprising amino acids 2 to 2455 of SEQ ID NO:16, wherein amino acids 2 to 2455 comprise a polypeptide of SEQ ID NO:16 minus the start methionine; and    (c) a polypeptide comprising amino acids 1 to 2455 of SEQ ID NO:16.    
     
     
         21 . The isolated polypeptide of  claim 20 , wherein the polypeptide comprises two or more sequential amino acid deletions from one or both of: 
 (a) the C-terminus of the polypeptide; and    (b) the N-terminus of the polypeptide.    
     
     
         22 . A method of identifying a compound that modulates the activity of the polypeptide of  claim 20 , the method comprising: 
 (a) determining the activity of the polypeptide of  claim 20  in the absence of a test compound;    (b) determining the activity of the polypeptide in the presence of a test compound; and    (c) comparing the activity of the polypeptide in the presence of the test compound with the activity of the polypeptide in the absence of the test compound,    wherein a change in the activity of the polypeptide in the presence of the test compound relative to the activity of the polypeptide in the absence of the test compound indicates that the compound that modulates the activity of the polypeptide.    
     
     
         23 . An isolated antibody which specifically binds to the polypeptide of  claim 20 .  
     
     
         24 . The antibody of  claim 23 , wherein the antibody is selected from the group consisting of a chimeric antibody, a single chain antibody, a Fab fragment, and a humanized antibody.  
     
     
         25 . A method of isolating an ACC polypeptide comprising: 
 (a) contacting crude lysate derived from a cell or tissue expressing an ACC polypeptide with an antibody to form a complex comprising an antibody and ACC;    (b) washing the complex with a buffer comprising 0.5 M NaCl; and    (c) contacting the complex with an eluting ligand.    
     
     
         26 . The method of  claim 25 , wherein the antibody comprises an IgG antibody.  
     
     
         27 . The method of  claim 26 , wherein the IgG antibody is a c-Myc-5 IgG antibody and the eluting ligand is a myc peptide.  
     
     
         28 . The method of  claim 27 , wherein the myc peptide comprises the amino acid sequence of SEQ ID NO:17.  
     
     
         29 . The method of  claim 26 , wherein the IgG antibody is an anti-FLAG IgG antibody.  
     
     
         30 . The method of  claim 25 , wherein the antibody is bound to a substrate.  
     
     
         31 . The method of  claim 25 , wherein the ACC polypeptide is an ACC1 polypeptide.  
     
     
         32 . The method of  claim 25 , wherein the ACC polypeptide is an ACC2 polypeptide.  
     
     
         33 . The method of  claim 25  wherein the ACC polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:13 and 16.  
     
     
         34 . A polynucleotide capable of inhibiting the expression of an ACC2 gene comprising a polynucleotide sequence selected from the group consisting of SEQ ID NOs:12 and 15 by antisense inhibition.  
     
     
         35 . A method of inhibiting ACC2 gene expression comprising introducing a polynucleotide of  claim 34  into a cell or tissue that expresses an ACC2 gene, thereby inhibiting the expression of the gene in the cell or tissue by antisense inhibition.  
     
     
         36 . A polynucleotide capable of inhibiting the expression of an ACC2 gene comprising a polynucleotide sequence selected from the group consisting of SEQ ID NOs:12 and 15 by RNA interference.  
     
     
         37 . A method of inhibiting ACC2 gene expression comprising introducing a polynucleotide of  claim 36  into a cell or tissue that expresses an ACC2 gene, thereby inhibiting the expression of the gene in the cell or tissue by RNA interference.  
     
     
         38 . An isolated polypeptide comprising an ACC2 polypeptide encoded by the cDNA deposited as ATCC Accession No. PTA-6054.  
     
     
         39 . A method of increasing the activity of a human ACC2 polypeptide, the method comprising generating an enhanced ACC2 polypeptide comprising 
 (a) a phenylalanine residue at position 254,    (b) a glutamine residue at position 346,    (c) a threonine residue at position 565,    (d) an asparagine at position 841,    (e) a valine residue at position 1103,    (f) a cysteine residue at position 1259,    (g) an alanine residue at position 1526, and    (h) an isoleucine residue at position 1717,    wherein the human ACC2 polypeptide does not comprise SEQ ID NO:13 and wherein the enhanced ACC2 polypeptide has an enzymatic activity level that is greater than the enzymatic activity level of an ACC2 polypeptide that does not contain the indicated residues at the indicated positions.    
     
     
         40 . The method of  claim 39 , wherein the human ACC2 polypeptide sequence is selected from the group consisting of SEQ ID NOs: 2, 4 and 6.