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  Acetyl coa carboxylase 2 sequences and methodsUSPTO Application #: 20080026363Title: 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. (end of abstract) Agent: Louis J. Wille Bristol-myers Squibb Company - Princeton, NJ, US Inventors: Dong Cheng, John N. Feder, Luping Chen, Ching-Hsuen Chu USPTO Applicaton #: 20080026363 - Class: 435004000 (USPTO) Related Patent Categories: Chemistry: Molecular Biology And Microbiology, Measuring Or Testing Process Involving Enzymes Or Micro-organisms; Composition Or Test Strip Therefore; Processes Of Forming Such Composition Or Test Strip The Patent Description & Claims data below is from USPTO Patent Application 20080026363. Brief Patent Description - Full Patent Description - Patent Application Claims
[0001] This application is a division of U.S. application Ser. No. 11/186,999, filed Jul. 21, 2005, which claims priority benefit to U.S. provisional application No. 60/590,948 filed Jul. 23, 2004. The entire teachings of the referenced application are incorporated herein by reference. FIELD OF THE INVENTION [0002] Disclosed and claimed herein are novel polynucleotides encoding human and rat Acetyl CoA Carboxylase 2 ("ACC2") polypeptides, fragments and homologues thereof. Vectors, host cells, antibodies, and recombinant and synthetic methods for producing the ACC2 polypeptides are provided. Further described are diagnostic and therapeutic methods for applying the ACC2 polypeptides to the diagnosis, treatment, and/or prevention of various diseases and/or disorders related to these polypeptides, including obesity. Further embodiments include screening methods for identifying agonists and antagonists of the polynucleotides and polypeptides of the present invention. BACKGROUND OF THE INVENTION [0003] Acetyl CoA carboxylase (ACC) is the rate-determining enzyme of fatty acid biosynthesis in plants and animals. ACC is a biotin containing enzyme which catalyzes the carboxylation of acetyl CoA to form malonyl CoA in a two-step reaction (Beaty & Lane, (1982). J. Biol. Chem. 257:924-929). The first step is the ATP-dependent carboxylation of biotin covalently linked to the enzyme. In the second step, a carboxyltransferase step, the carboxyl group is transferred to the substrate, acetyl CoA, to form malonyl CoA. Citrate is a potent allosteric activator of ACC. Malonyl CoA is the C2 donor for de novo synthesis of long chain fatty acids. [0004] In mammals, there are two subtypes of ACC, ACC1 and ACC2. ACC1 is mainly localized in lipogenic tissues such as adipose tissue and liver, where fatty acids are synthesized. ACC2 is found primarily in non-lipogenic tissues such as skeletal muscle and heart muscle, although some is also found in liver. Malonyl CoA allosterically inhibits carnitine palmitoyl transferase 1 (CPT1), which is a critical enzyme to transfer the long chain fatty acid into the mitochondria for .beta.-oxidation. Because ACC2 is co-localized with CPT-1, the primary role of malonyl CoA that is synthesized by ACC2 has been suggested to regulate the rate of .beta.-oxidation. [0005] ACC is a potential target in metabolic diseases for the treatment of metabolic syndrome including obesity, insulin resistance and dyslipidemia. Increased rates of muscle fatty acid oxidation, a reduced fat content and a reduction in total body fat were observed in ACC-2 knock-out mice (Abu-Elheiga et al., (2001) Science 291:2613-2616; Abu-Elheiga et al., (2003) Proc. Natl. Acad. Sci. USA. 100:10207-10212). Harwood et al. reported that ACC inhibitors caused reduction in fatty acid synthesis, increase in fatty acid oxidation, and reduction of respiratory quotient in rats (Harwood et al., (2003) J. Biol. Chem. 278:37099-37111). Chronic dosing of these compounds resulted in the reduction of whole body fat mass and improvement of insulin sensitivity (Harwood et al., (2003) J. Biol. Chem. 278:37099-37111). These observations further validated the enzyme as a drug target. [0006] Several human ACC2 and rat ACC2 nucleotide and amino acid sequences have been published (see, e.g., Human ACC2: GenBank Accession No. NM.sub.--001093 (SEQ ID NOs:1 and 2) and GenBank Accession No. AC007637 (SEQ ID NOs:3 and 4); Rat ACC2: GenBank Accession No. NM.sub.--053922 (SEQ ID NOs:7 and 8) and GenBank Accession No. AB004329 (SEQ ID NOs:9 and 10)). It was found, however, that for each species, each of the published amino acid and/or nucleotide sequences was different from one another by one or more residues. More specifically, it was found that the nucleotide sequences of human ACC2 and rat ACC2 contain non-silent mutations that introduce substitutions into several of the published encoded amino acid sequences of these enzymes. [0007] In order to identify the most effective modulators of human ACC2 and rat ACC2, accurate nucleotide and amino acid sequences are required. Therefore, what is needed to advance research on human and rat ACC2 is an accurate amino acid sequence for these enzymes, as well as the encoding nucleotide sequences. The subject matter disclosed and claimed herein solves this and other problems. SUMMARY OF THE INVENTION [0008] Described and claimed herein is an isolated nucleic acid molecule encoding a human ACC2 polypeptide. In one embodiment the nucleic acid molecule comprises a polynucleotide having a nucleotide sequence selected from the group consisting of: (a) a polynucleotide encoding an ACC2 polypeptide comprising SEQ ID NO:12; (b) an isolated polynucleotide encoding a human ACC2 polypeptide comprising amino acids 2 to 2458 of SEQ ID NO:12 minus the start methionine; (c) an isolated polynucleotide encoding a human ACC2 polypeptide comprising amino acids 1 to 2458 of SEQ ID NO:12 including the start codon; (d) an isolated polynucleotide encoding the ACC2 polypeptide encoded by the cDNA clone contained in 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. The isolated nucleic acid molecule can comprise, for example, the nucleotide sequence of SEQ ID NO:11. In additional aspects, the present invention also relates to a polynucleotide that is complementary to the isolated nucleic acid molecule, a vector comprising the isolated nucleic acid molecule and a host cell, which can be a mammalian host cell, comprising the vector. [0009] Also disclosed is a method of making an isolated human ACC2 polypeptide. In one embodiment the method comprises: (a) culturing the recombinant host cell under conditions such that the polypeptide is expressed; and (b) recovering the polypeptide. [0010] Further described is an isolated human ACC2 polypeptide. In one embodiment the polypeptide comprises an amino acid sequence selected from the group consisting of: (a) a polypeptide comprising SEQ ID NO:12; (b) a polypeptide comprising amino acids 2 to 2458 of SEQ ID NO:12, wherein amino acids 2 to 2458 comprise a polypeptide of SEQ ID NO:12 minus the start methionine; and (c) a polypeptide comprising amino acids 1 to 2458 of SEQ ID NO:12. In another embodiment, 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.sub.2-terminus of the polypeptide. [0011] Another embodiment disclosed herein is a method of identifying a compound that modulates the activity of a human ACC2 polypeptide. In one embodiment, the method comprises: (a) determining the activity of an ACC2 polypeptide 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. [0012] A further embodiment is an isolated antibody which specifically binds to a human ACC2 polypeptide. In various embodiments, the antibody is selected from the group consisting of a chimeric antibody, a single chain antibody, a Fab fragment, and a humanized antibody. [0013] Further disclosed and claimed is an isolated nucleic acid molecule encoding a rat ACC2 polypeptide. In one embodiment the isolated nucleic acid molecule comprises a polynucleotide having a nucleotide sequence selected from the group consisting of: (a) a polynucleotide encoding an ACC2 polypeptide comprising SEQ ID NO:14; (b) an isolated polynucleotide encoding a rat ACC2 polypeptide comprising amino acids 2 to 2458 of SEQ ID NO:14 minus the start methionine; (c) an isolated polynucleotide encoding a rat ACC2 polypeptide comprising amino acids 1 to 2458 of SEQ ID NO:14 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. [0014] In one embodiment, the isolated rat nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:13. In additional aspects, the present invention comprises a polynucleotide that is complementary to the isolated nucleic acid molecule, a vector comprising the isolated nucleic acid molecule and a host cell, which can be a mammalian host cell, comprising the vector. [0015] Another embodiment relates to a method of making an isolated rat ACC2 polypeptide. In one embodiment the method comprises: (a) culturing a recombinant host cell under conditions such that the polypeptide is expressed; and (b) recovering the polypeptide. [0016] Yet a further embodiment is an isolated rat ACC2 polypeptide. For example, the polypeptide comprises an amino acid sequence selected from the group consisting of: (a) a polypeptide comprising SEQ ID NO:14; (b) a polypeptide comprising amino acids 2 to 2455 of SEQ ID NO:14, wherein amino acids 2 to 2455 comprise a polypeptide of SEQ ID NO:14 minus the start methionine; and (c) a polypeptide comprising amino acids 1 to 2455 of SEQ ID NO:14. In another embodiment, 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.sub.2-terminus of the polypeptide. [0017] Further disclosed is a method of identifying a compound that modulates the activity of a rat ACC2 polypeptide. In one embodiment, the method comprises: (a) determining the activity of a polypeptide 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. [0018] An isolated antibody that specifically binds to the rat ACC2 polypeptide is disclosed. In various embodiments, the antibody is selected from the group consisting of a chimeric antibody, a single chain antibody, a Fab fragment, and a humanized antibody. [0019] A method of isolating a rat ACC polypeptide is additionally disclosed. In one embodiment, the method comprises: (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 an ACC; (b) washing the complex with a buffer comprising 0.5 M NaCl; and (c) contacting the complex with an eluting ligand. The antibody can comprise an IgG antibody, and in one embodiment, can be, for example, a c-Myc-5 IgG antibody and the eluting ligand can be a myc peptide. The myc peptide can comprise the amino acid sequence of SEQ ID NO:16. In another example, the IgG antibody is an anti-FLAG IgG antibody. Further, the antibody can be bound to a substrate. The method can be employed to isolate any ACC polypeptide, such as an ACC1 or ACC2 polypeptide. [0020] Additionally, a polynucleotide capable of inhibiting the expression of an ACC2 gene comprising a polynucleotide sequence selected from the group consisting of SEQ ID NOs:11 and 13 by antisense inhibition is disclosed, as well as a method of inhibiting ACC2 gene expression comprising introducing an antisense polynucleotide 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. [0021] A polynucleotide capable of inhibiting the expression of an ACC2 gene comprising a polynucleotide sequence selected from the group consisting of SEQ ID NOs:11 and 13 by RNA inhibition is disclosed, as well as a method of inhibiting ACC2 gene expression comprising introducing an RNAi polynucleotide into a cell or tissue that expresses an ACC2 gene, thereby inhibiting the expression of the gene in the cell or tissue by RNA inhibition. Continue reading... Full patent description for Acetyl coa carboxylase 2 sequences and methods Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Acetyl coa carboxylase 2 sequences and methods patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. 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