A Hypersweet Protein: Removal of The Specific Negative Charge at Asp21 Enhances Thaumatin Sweetness.
A Hypersweet Protein: Removal of The Specific Negative Charge at Asp21 Enhances Thaumatin Sweetness.
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DOI：
10.1038/srep20255
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发表时间：
2016-02-03
影响因子：
4.6
通讯作者：
Kitabatake N
中科院分区：
文献类型：
--
作者：
Masuda T;Ohta K;Ojiro N;Murata K;Mikami B;Tani F;Temussi PA;Kitabatake N
奇异果甜蛋白是一种强烈的甜味蛋白质，在50 nM的浓度下可以增强甜味，该浓度比蔗糖的摩尔浓度大100，000倍。在这里，我们试图通过去除索马甜与味觉受体相互作用一侧的负电荷来生产具有增强甜味的蛋白质。我们获得了一个D21 N突变体，其阈值为31 nM，比野生型索马甜甜得多，并且与单链莫内林的Y 65 R突变体一起，是迄今为止已知的两种最甜的蛋白质之一。T1 R2-T1 R3甜味受体和奇异果甜蛋白之间的复合物模型，来源于楔形模型框架中的栓系对接，证实了每个对甜味至关重要的带正电荷的残基接近相反电荷的受体残基，以产生最佳的静电相互作用。此外，D21和其可能的对应物D433（位于受体的T1 R2原聚体上）之间的距离安全地大以避免静电排斥，但同时，如果D21突变成相应的天冬酰胺，则可以更接近。这些发现清楚地证实了静电势在索马甜与甜味受体相互作用中的重要性。
Thaumatin is an intensely sweet-tasting protein that elicits sweet taste at a concentration of 50 nM, a value 100,000 times larger than that of sucrose on a molar basis. Here we attempted to produce a protein with enhanced sweetness by removing negative charges on the interacting side of thaumatin with the taste receptor. We obtained a D21N mutant which, with a threshold value 31 nM is much sweeter than wild type thaumatin and, together with the Y65R mutant of single chain monellin, one of the two sweetest proteins known so far. The complex model between the T1R2-T1R3 sweet receptor and thaumatin, derived from tethered docking in the framework of the wedge model, confirmed that each of the positively charged residues critical for sweetness is close to a receptor residue of opposite charge to yield optimal electrostatic interaction. Furthermore, the distance between D21 and its possible counterpart D433 (located on the T1R2 protomer of the receptor) is safely large to avoid electrostatic repulsion but, at the same time, amenable to a closer approach if D21 is mutated into the corresponding asparagine. These findings clearly confirm the importance of electrostatic potentials in the interaction of thaumatin with the sweet receptor.