Western blot analysis of MCF7 cells, untreated (-) or treated with Adenosine-2', 3'-dialdehyde (AdOx, 100 μM, 24 hr; +), using Asymmetric Di-Methyl Arginine Motif [adme-R] MultiMab™ Rabbit mAb mix (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
The specificity of Asymmetric Di-Methyl Arginine Motif [adme-R] MultiMab™ Rabbit mAb mix was determined using peptide ELISA. The figure demonstrates that the antibody is specific for asymmetric di-methyl arginine and does not react with mono-methyl, di-methyl or tri-methyl lysine and does not react with mono-methyl or symmetric di-methyl arginine.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
Asymmetric Di-Methyl Arginine Motif [adme-R] MultiMab™ Rabbit mAb mix recognizes endogenous levels of proteins that are asymmetrically dimethylated on arginine residues. This antibody does not cross-react with monomethylated, symmetrically methylated arginine, or methylated lysine residues.
All Species Expected
MultiMab™ rabbit monoclonal mix antibodies are prepared by combining individual rabbit monoclonal clones in optimized ratios for the approved applications. Each antibody in the mix is carefully selected based on motif recognition and performance in multiple assays. Each mix is engineered to yield the broadest possible coverage of the modification being studied while ensuring a high degree of specificity for the modification or motif.
Arginine methylation is a prevalent PTM found on both nuclear and cytoplasmic proteins. Arginine methylated proteins are involved in many different cellular processes, including transcriptional regulation, signal transduction, RNA metabolism, and DNA damage repair (1-3). Arginine methylation is carried out by the arginine N-methyltransferase (PRMT) family of enzymes that catalyze the transfer of a methyl group from S-adenosylmethionine (AdoMet) to a guanidine nitrogen of arginine (4). There are three different types of arginine methylation: asymmetric dimethylarginine (aDMA, omega-NG,NG-dimethylarginine), where two methyl groups are placed on one of the terminal nitrogen atoms of the guanidine group of arginine; symmetric dimethylarginine (sDMA, omega-NG,N’G-dimethylarginine), where one methyl group is placed on each of the two terminal guanidine nitrogens of arginine; and monomethylarginine (MMA, omega-NG-dimethylarginine), where a single methyl group is placed on one of the terminal nitrogen atoms of arginine. Each of these modifications has potentially different functional consequences. Though all PRMT proteins catalyze the formation of MMA, Type I PRMTs (PRMT1, 3, 4, and 6) add an additional methyl group to produce aDMA, while Type II PRMTs (PRMT5 and 7) produce sDMA. Methylated arginine residues often reside in glycine-arginine rich (GAR) protein domains, such as RGG, RG, and RXR repeats (5). However, PRMT4/CARM1 and PRMT5 methylate arginine residues within proline-glycine-methionine rich (PGM) motifs (6).
Symmetrically dimethylated (sDMA) histone H4R3 is prevalent in undifferentiated mouse embryonic neural precursors, but both symmetric and asymmetric dimethyl (aDMA) H4R3 modifications are detected in post-mitotic neurons and developing oligodendrocytes during later stages of development. This implies that sDMA modifications may be negative epigenetic regulatory events while aDMA modifications may signal epigenetic activation sites (7).
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