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G protein coupled acceptor protein, its production method and use

A technology for coupling receptors and G proteins, applied in the field of new G protein-coupled receptor proteins and new G protein-coupled receptor proteins, can solve the problems of DNA discovery of receptor proteins that have not been reported

Inactive Publication Date: 2001-11-14
TAKEDA PHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the case of galanin and amylin, no findings regarding their receptor protein DNA have been reported

Method used

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  • G protein coupled acceptor protein, its production method and use
  • G protein coupled acceptor protein, its production method and use
  • G protein coupled acceptor protein, its production method and use

Examples

Experimental program
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Embodiment approach

[0641] Particularly preferred embodiments of the invention include:

[0642] 8. A method for screening DNA encoding the amino acid sequence of a G protein-coupled receptor protein from a DNA library, the method comprising performing a polymerase chain reaction in the presence of a mixture of the following substances to selectively amplify the encoded G protein contained in said DNA library Say the DNA of the amino acid sequence of the G protein-coupled receptor protein:

[0643] (1) said DNA library,

[0644] (2) at least one DNA primer selected from DNA primers having a nucleotide sequence represented by SEQ ID NO: 1, and

[0645] (3) at least one DNA primer selected from DNA primers having a nucleotide sequence represented by SEQ ID NO:2;

[0646] 9. A method for screening DNA encoding the amino acid sequence of a G protein-coupled receptor protein from a DNA library, the method comprising performing a polymerase chain reaction in the presence of a mixture of the f...

Embodiment 1

[1008] Preparation of synthetic DNA for amplifying DNA encoding G protein-coupled receptor proteins

[1009] Comparison of deoxyribonucleotide sequences encoding the known amino acid sequences corresponding to or close to the first transmembrane domains of the following proteins: human-derived TRH receptor protein (HTRHR), human-derived RANTES receptor protein ( L10918, HUMRANTES), (from human Burkitt's lymphoma unknown ligand receptor protein (X68149, HSBLR1A), (from human somatostatin receptor protein L14856, HUMSOMAT), from rat μ-opioid Substance receptor protein (U02083, RNU02083), rat-derived κ-opioid receptor protein (U00442), U00442, human-derived neuromedin B receptor protein (M73482, HUMNMBR), human-derived Muscarinic acetylcholine receptor protein (X15266, HSHM4), epinephrine α from rat 1 B receptor protein (L08609, RATAADREO1), human-derived somatostatin 3 receptor protein (M96738, HUMSSTR3X), human-derived C 5 a receptor protein (HUMCSAAR), human-derived unknown ...

Embodiment 2

[1021] Example 2 Isolation of DNA encoding human somatostatin receptor protein, DNA encoding human D5 dopamine receptor protein, and DNA encoding rat somatostatin receptor protein

[1022] (1) Amplify DNA by polymerase chain reaction (PCR) Use 1 ng each of cDNA (Quickclone, CLONTECH Laboratories) prepared from human brain tonsil, human pituitary gland and rat brain as a template, and 1 μM each of the DNA primers prepared in Example 1 , 2.5 mM dNTPs (deoxyribonucleoside triphosphate) and 2.5 units of TaqDNA polymerase (Takara Shuzo, Japan) were mixed with the buffer attached to the enzyme kit to make a total of 100 µl. The polymerase chain reaction was performed using a thermocycler manufactured by Perkin-Elmer. Set up a cycle consisting of 96°C for 30 seconds, 45°C for 1 minute and 60°C for 3 minutes. In total this cycle was repeated 30 times to amplify the DNA. Amplification of DNA was confirmed by 1.2% agarose gel electrophoresis [Fig. 17].

[1023] (2) S...

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Abstract

The present invention provides DNA primers effective in screening G protein coupled receptor protein-encoding DNA fragments. The primers which are complementary to nucleotide sequences that are in community with (homologous to) the nucleotide sequences encoding amino acid sequences corresponding to or near the first membrane-spanning domain or the sixth membrane-spanning domain each of known various G protein coupled receptor proteins were designed and synthesized. Methods of amplifying G protein coupled receptor protein-encoding DNAs using the above DNA primers, and novel target G protein coupled receptor protein-encoding DNAs are also provided. Screening of DNA libraries can be efficiently carried out. The invention also provides human pituitary gland or amygdala-derived and mouse pancreas-derived G protein coupled receptor proteins, etc. or salts thereof, partial peptides thereof.

Description

[0001] Technical Field of the Invention [0002] The present invention relates to novel DNA useful as a DNA primer for polymerase chain reaction (PCR); a method for amplifying various DNAs encoding G protein-coupled receptor proteins by PCR using the DNA primer; Method for screening various DNAs encoding G protein-coupled receptor proteins; DNA encoding G protein-coupled receptor proteins obtained by said screening method; G protein-coupled proteins encoded by DNA obtained by said screening method Receptor proteins, peptide fragments or segments thereof, and modified peptide derivatives thereof; and the like. [0003] The present invention also relates to novel G protein-coupled receptor proteins; DNA encoding novel G protein-coupled receptor proteins; methods of producing said G protein-coupled receptor proteins; said receptor proteins and proteins encoding said proteins Use of DNA; etc. [0004] The present invention also relates to novel G protein-coupled rec...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/705C07K14/72C12N15/12C12Q1/68
CPCC07K14/70571C07K14/723C12Q1/6876C07K14/705C12Q2600/158
Inventor 日沼州司细谷昌树藤井亮大泷彻也福住昌司大仪和宏
Owner TAKEDA PHARMA CO LTD
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