Amines with N-Acetyl Homocysteine Thiolactone

Modification of Amines with N-Acetyl Homocysteine Thiolactone 

Thiolation of peptides and other small molecules containing amines proceeds easily with N-acetyl homocysteine thiolactone. However, protein modification often results in much lower yields unless the reaction is done for extended periods at pH 10-11. 

It has been found that silver ions catalyze the thiolation process with proteins, allowing the reaction to be completed rapidly at physiological pH (Benesch and Benesch, 1958). The addition of an equal molar concentration of AgNO j forms an insoluble complex with the thiolactone, and this in turn reacts with protein amines. 

Dissolve the amine-containing molecule to be thiolated at a concentration of lOmg/ml in cold (4°C) 1M sodium bicarbonate (reaction buffer). For proteins, dissolve them in deionized water at a pH of 7.0-7.5, at room temperature. Note The presence of some buffer salts, like phosphate or carbonate, is incompatible with silver nitrate. 

Add N-acetyl homocysteine thiolactone (Aldrich) to the bicarbonate reaction mixture to obtain a concentration representing a 10- to 20-fold excess over the amount of amines present. For protein thiolation, add the same molar excess of thiolactone reagent to the water reaction medium, and then slowly add an equivalent molar quantity of silver nitrate (AgNO j). Maintain the pH at 7.0-7.5 with periodic addition of NaOH. 

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