A novel way to generate nonspecifically radiolabeled proteins for use as molecular-weight markers.
A method for labeling proteins using radioactive nucleotides in the presence of divalent cations such as Cu2+, Zn2+ and Cd2+ is described. Small amounts of marker proteins can be rapidly labeled using the method and they can be used for molecular-weight determination of radioactive proteins of unknown molecular weights, as for example, any protein labeled with 35S-methionine in vivo or any protein radiolabeled by 32P in protein phosphorylation experiments. The gels in which the labeled markers and the proteins of unknown molecular weight are electrophoresed could be directly exposed to X-ray films and comparisons made from a single autoradiogram, avoiding a two step procedure of Coomassie blue staining followed by autoradiography and aligning of the two sets of bands. This metal-ion-mediated labeling of proteins described in the present communication is not by any site-specific interaction of the nucleotide with the proteins and the protein is denatured after the complex formation. The adducts, once formed, are stable under conditions of SDS-PAGE or EDTA treatment. It is suggested that proteins labeled under conditions described in this communication are ternary complexes involving aromatic residues of the proteins, nucleotides and the divalent cations.
Mohanty
B K
BK
Centre for Cellular and Molecular Biology, Hyderabad, India.
Krishna
B M
BM
Das
M R
MR
D002104:Cadmium / Q000737:chemistry; D003300:Copper / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007553:Isotope Labeling / Q000379:methods*; D008970:Molecular Weight; D009711:Nucleotides / Q000737:chemistry; D010455:Peptides / Q000138:chemical synthesis; D010761:Phosphorus Radioisotopes; D011506:Proteins / Q000138:chemical synthesis*; D012680:Sensitivity and Specificity; D013696:Temperature; D014316:Tritium; D015032:Zinc / Q000737:chemistry
D009711:Nucleotides; D010455:Peptides; D010761:Phosphorus Radioisotopes; D011506:Proteins; D002104:Cadmium; D014316:Tritium; D003300:Copper; D015032:Zinc