• Mildred Elley - NYC Campus

• QWER 12345

UREA Product Numbers: U6504, Electrophoresis Reagent U5378, Molecular Biology Reagent U4128, Cell Culture Tested U0631, SigmaUltra U4884, USP U5128, ACS Reagent U1250, Pellets Storage Temperature Room Temperature Cas #: 57-13-6 Synonyms: Carbamide, Carbonyl diamide Product Description Appearance: U4128 and U1250 are white pellets; others are white powder Molecular Formula: CH4N2O Molecular Weight: 60.06 Urea is a mild chaotropic agent useful for the solubilization and denaturation of proteins.1-3 It is also useful for renaturing proteins from samples already denatured with 6 M guanidine hydrocholride such as inclusion bodies.4,5 It is commonly used to solubilize and denature proteins for denaturing isoelectric focusing and two-dimensional electrophoresis.2,3,6,7,8 Preparing samples for two-dimensional polyacrylamide electrophoresis (2D PAGE) involves solubilization, denaturation and reduction in order to completely disrupt the interactions between the proteins.2 Due to the diversity of samples which are analyzed by 2D gel electrophoresis, there is no single method of sample preparation that works in all cases. Ideally, all noncovalently bound protein complexes and aggregates should be denatured to form a solution of individual polypeptides.3 However, regardless of the method used, protein modifications which might result in artifactual spots on 2D maps must be minimized. For example, samples containing urea must not be heated as this may introduce considerable charge heterogeneity due to carbamylation of the proteins (see information below). Samples should kept cold at all times and handling should be kept to a minimum. Urea in solution is in equilibrium with ammonium cyanate. The form that reacts with protein amino groups is isocyanic acid. Urea in the presence of heat and protein leads to carbamylation of the proteins. Carbamylation by isocyanic acid interferes with protein characterization because isocyanic acid reacts with the amino terminus of proteins, preventing N-terminal sequencing. Isocyanic acid also reacts with side chains of lysine and arginine residues resulting in a protein that is unsuitable for many enzymatic digests. In addition, carbamylation often leads to confusing results from peptides having unexpected retention times and masses.9 When performing enzymatic protein digests it is important to remove urea first. Even though some enzymes will tolerate small amounts of urea, the elevated temperature used for most reactions will lead to carbamylation during the course of the digest. The urea can be removed prior to digest by fast reversed phase chromatography, spin columns, or dialysis. Urea is used in cell or tissue culture media to increase the osmolality.10 It is the principal end product of nitrogen metabolism in most mammals, formed by the enzymatic reactions of the Kreb’s cycle. Preparation Instructions One gram of urea dissolves in one ml water, 10 ml 95% alcohol, 20 ml absolut