Claims
- 1. A method for identifying the structure of a dimethylarginine comprising:
(i) obtaining, by mass spectroscopy, a neutral loss spectra of a peptide containing a dimethylarginine, and (ii) ascertaining if the neutral loss spectra shows one or both of neutral loss of monomethylamine (MMA), dimethylcarbodiimide (DMC), and/or neutral loss of dimethylamine (DMA), wherein neutral loss of monomethylamine (MMA) and of dimethylcarbodiimide (DMC) indicates the presence of a symmetrically dimethylated arginine residue, whereas neutral loss of dimethylamine (DMA) indicates the presence of a asymmetrically dimethylated arginine residue.
- 2. A method for identifying dimethylarginine residues in a test peptide, comprising:
(i) identifying the presence of dimethylarginine residues from mass spectra of a sample peptide obtained under conditions in which the spectra reveal mass modification by methylation, if any, of arginine residue in the sample peptide; (ii) ascertaining the nature of the methylation of a dimethylarginine residue identifying in step (i) by determining if a neutral loss spectra of the sample peptide shows one or both of neutral loss of monomethylamine (MMA), dimethylcarbodiimide (DMC), and/or neutral loss of dimethylamine (DMA), wherein neutral loss of monomethylamine (MMA) and of dimethylcarbodiimide (DMC) indicates the presence of a symmetrically dimethylated arginine residue, whereas neutral loss of dimethylamine (DMA) indicates the presence of a asymmetrically dimethylated arginine residue.
- 3. A method for identifying substrates of a protein arginine methyltransferase comprising:
(i) obtaining mass spectra for one or more sample polypeptides which have been exposed to a protein arginine methyltransferase (PRMT) under conditions wherein methylation, if any, of arginine residues in a substrate protein of the PRMT can occur; (ii) identifying the presence of dimethylarginine residues in the sample polypeptide by the presence of mass modified arginine in the mass spectra; and (iii) ascertaining the nature of the methylation of a dimethylarginine residue identifying in step (i) by determining if a neutral loss spectra of the sample peptide shows one or both of neutral loss of monomethylamine (MMA), dimethylcarbodiimide (DMC), and/or neutral loss of dimethylamine (DMA), wherein neutral loss of monomethylamine (MMA) and of dimethylcarbodiimide (DMC) indicates the presence of a symmetrically dimethylated arginine residue whereas neutral loss of dimethylamine (DMA) indicates the presence of a asymmetrically dimethylated arginine residue.
- 4. The method of claim 3, further comprising determining the substrate specificity of a protein arginine methyltransferase by determining the sequence of at least that portion of a polypeptide including a dimethylarginine residue.
- 5. The method of claim 3, wherein said one or more sample polypeptides constitute a library of sample polypeptides.
- 6. The method of any of claims 2-5, wherein the neutral loss spectra is generated from said mass spectra.
- 7. The method of any of claims 1-5, wherein the spectra are obtained using a mass spectrometer in which ionization of the sample protein is accomplished by matrix-assisted laser desorption (MALDI) ionization, electrospray (ESI), or electron impact (EI).
- 8. The method of any of claims 1-5, wherein the spectra are obtained using ESI-MS/MS.
- 9. A mass spectrometry system including an automated system for identifying dimethylarginine residues in a test peptide, which automated system comprises:
(i) a methylation detection system that (a) analyzes one or more mass spectra obtained under conditions in which the spectra reveal mass modification by methylation, if any, of arginine residue in a sample peptide, and (b) identifies the presence of dimethylarginine residues from characteristic mass differences between arginine and dimethylarginine in the mass spectra; (ii) a neutral loss detection system for determining the nature of the methylation of a dimethylarginine residue identified by the methylation detection system, which neutral loss detection system determines if a neutral loss spectra of the sample peptide shows one or both of neutral loss of monomethylamine (MMA), dimethylcarbodiimide (DMC), and/or neutral loss of dimethylamine (DMA), wherein neutral loss of monomethylamine (MMA) and of dimethylcarbodiimide (DMC) indicates the presence of a symmetrically dimethylated arginine residue, whereas neutral loss of dimethylamine (DMA) indicates the presence of a asymmetrically dimethylated arginine residue.
- 10. A method of conducting a drug discovery business, comprising:
(i) by the method of any of claims 1-8, determining the identity of a PRMT and substrate thereof; (ii) identifying agents by their ability to alter the level of methylation of the substrate; (iii) conducting therapeutic profiling of agents identified in step (ii), or further analogs thereof, for efficacy and toxicity in animals; and (iv) formulating a pharmaceutical preparation including one or more agents identified in step (iii) as having an acceptable therapeutic profile.
- 11. The method of claim 10, including an additional step of establishing a distribution system for distributing the pharmaceutical preparation for sale, and may optionally include establishing a sales group for marketing the pharmaceutical preparation.
- 12. A method of conducting a proteomics business, comprising:
(i) by the method of any of claims 1-8, determining the identity of a PRMT and substrate thereof; (ii) licensing, to a third party, the rights for further drug development of agents that alter the level of methylation of the substrate.
- 13. A protein sequence or coding sequence database comprising annotation information representative of the methylation status of arginine residues.
REFERENCE TO RELATED APPLICATION
[0001] This Application claims priority to U.S. Provisional Application 60/292,075, filed on May 18, 2001, the entire content of which is hereby incorporated by reference.
Provisional Applications (1)
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Number |
Date |
Country |
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60292075 |
May 2001 |
US |