Cagan R H, Morris J A
Proc Soc Exp Biol Med. 1976 Sep;152(4):635-40. doi: 10.3181/00379727-152-39457.
The presence of a single cysteine in the sweet-tasting protein monellin was confirmed by titrations with p-hydroxymercuribenzoate (PHMB) and 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB). The sulfhydryl group in native monellin reacts very slowly with each of these reagents, indicating that the sulfhydryl is relatively inaccessible. In the presence of either 6 M guanidine-HCl, 8 M urea, or 1% sodium dodecyl sulfate, the rate of reaction of the sulfhydryl group with titrant is dramatically increased. Under a variety of conditions, the presence of 1 mole of sulfhydryl per mole of protein (of molecular weight 10,700) was found. Reaction of the sulfhydryl by titration with PHMB or DTNB leads to loss of sweetness. The free sulfhydryl is also lost by carboxymethylation of monellin in the presence of guanidine-HCl, yielding a protein that is not sweet. Exposure to air in the presence of denaturant leads to a decrease in the sweetness of monellin. Sweetness of the PHMB-reacted monellin can be recovered upon treatment of the protein with mercaptoethanol, and the partial loss of sweetness that occurs with air exposure is lessened in the presence of mercaptoethanol. It is postulated that alteration of the single sulfhydryl group of monellin leads to a change in the tertiary structure of the protein and hence its sweet taste.
通过用对羟基汞苯甲酸（PHMB）和5,5'-二硫代双（2-硝基苯甲酸）（DTNB）进行滴定，证实了甜味蛋白莫内林中存在单个半胱氨酸。天然莫内林中的巯基与这些试剂中的每一种反应都非常缓慢，这表明该巯基相对难以接近。在6 M盐酸胍、8 M尿素或1%十二烷基硫酸钠存在的情况下，巯基与滴定剂的反应速率会显著增加。在各种条件下，发现每摩尔蛋白质（分子量为10700）含有1摩尔巯基。用PHMB或DTNB滴定巯基会导致甜味丧失。在盐酸胍存在的情况下，通过对莫内林进行羧甲基化也会使游离巯基丧失，产生一种不甜的蛋白质。在变性剂存在的情况下暴露于空气中会导致莫内林的甜度降低。用巯基乙醇处理蛋白质后，PHMB反应后的莫内林的甜度可以恢复，并且在巯基乙醇存在的情况下，因暴露于空气中而导致的部分甜度损失会减少。据推测，莫内林单个巯基的改变会导致蛋白质三级结构的变化，从而导致其甜味发生改变。
