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Dithiothreitol (DTT), CAS 3483-12-3 a new type of green additive

Dithiothreitol (DTT) is a commonly used reducing agent, also known as the new green additive. It is a small molecular organic compound with two mercaptan groups (-SH). Due to its reducing properties and stability, DTT is widely used in biochemistry and molecular biology experiments.

The main role of DTT is to reduce disulfide bonds in proteins and other biomolecules. The disulfide bond is an important part of protein folding and stability, but under certain experimental conditions, such as reducible SDS-PAGE analysis, protein recombination and folding, it is necessary to reduce the disulfide bond to two thiol groups to unravel the spatial structure of the protein. DTT can react with disulfide bonds to reduce them to mercaptan groups, thus opening up the spatial structure of the protein and making it easy to analyze and manipulate.

DTT can also be used to protect enzyme activity and stability. In some enzyme-catalyzed reactions, the activity of the enzyme may be reduced by the oxidant. DTT can react with oxidants to reduce them to harmless substances, thereby protecting the activity and stability of the enzyme.

Compared to traditional reducing agents such as β-mercaptoethanol (β-ME), DTT is considered to be a safer and more stable reducing agent. It is not only stable in aqueous solution, but also maintains its reducing properties under high temperature and acid-base conditions.

The use of DTT is relatively simple. In general, DTT is dissolved in an appropriate buffer and then added to the experimental system. The optimal concentration of DTT needs to be determined according to the specific experiment, and is generally used in the range of 0.1-1mM. Lower concentrations can reduce adverse effects on cell growth and can reduce cytotoxicity due to overexpression of target proteins. Higher concentrations may cause excessive cell metabolic burden, affecting cell growth and expression efficiency.

The way to determine the optimal concentration can be to evaluate the expression level of the target protein by conducting IPTG induction tests at different concentrations. Small-scale culture tests can be performed using a range of IPTG concentrations (e.g. 0.1 mM, 0.5 mM, 1 mM, etc.) and the expression effect at different concentrations can be evaluated by detecting the expression level of the target protein (e.g. Western blot or fluorescence detection). According to the experimental results, the concentration with the best expression effect was selected as the optimal concentration.

In addition, you can also refer to the relevant literature or the experience of other laboratories to understand the commonly used IPTG concentration range under similar experimental conditions, and then optimize and adjust according to the experimental needs.

It is important to note that the optimal concentration may vary depending on different expression systems, target proteins, and experimental conditions, so it is best to optimize on a case-by-case basis.

In summary, DTT is a commonly used reducing agent that can be used to reduce disulfide bonds in proteins and other biomolecules and to protect enzyme activity and stability. It has been widely used in biochemistry and molecular biology experiments.