DL-Dithiothreitol

Dithiothreitol (DTT) is a disulfide (SH) reducing agent found in proteins and in the amino acid cysteine. It reduces a disulfide bond by forming a six-membrane ring and initiating a thiol-disulfide interchange. It is almost 7-fold stronger reducing agent than βME, hence efficient in lower concentrations.

An excellent reagent for maintaining SH groups in reduced state; quantitatively reduces disulfides. DTT is effective in sample buffers for reducing protein disulfide bonds prior to SDS-PAGE. DTT can also be used for reducing the disulfide bridge of the cross-linker N,N′-bis(acryloyl)cystamine to break apart the matrix of a polyacrylamide gel. DTT is less pungent and is less toxic than 2-mercaptoethanol. Typically, a seven fold lower concentration of DTT (100 mM) is needed than is used for 2-mercaptoethanol (5% v/v, 700 mM).

DTT has been used:

• as one of the reactants in the reduction and alkylation of αs1-Casein, the major allergen of cow′s milk.[1]
• as a component of medium for the demembranation and reactivation of spermatozoa.[2]
• to maintain stability of the enzyme as thiol effectively protects the active sites of the biocatalyst.[3]
• as a reducing agent to test the specificity of the reaction of N-Ethylmaleimide with sulfhydryl groups.[4]
• in proteomics analysis as in-solution protein digestion for mass spectrometry[5]
• as a buffer component for protein quantification, to prepare wash buffer, lysis buffer, sample buffer, and protein elution buffer[6][4]

Dithiothreitol (DTT) is extensively applied in chemical peptide synthesis and biochemical preparations of thiol proteins. It is also used in protein chemistry studies, such as protein folding and enzyme activity. It also acts as an enzyme stabilizer to stop DNA modification. DTT specifically mediates the thiol-disulfide interchange reaction to entirely reduce the intra- or inter-molecular disulfide bonds in biomolecules. This reaction results in the formation of thiols and the cyclic disulfide of DTT.[7]

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