WHY DO SOME ORGANISMS USE A UREA-METHYLAMINE MIXTURE AS OSMOLYTE - THERMODYNAMIC COMPENSATION OF UREA AND TRIMETHYLAMINE N-OXIDE INTERACTIONS WITH PROTEIN
WHY DO SOME ORGANISMS USE A UREA-METHYLAMINE MIXTURE AS OSMOLYTE - THERMODYNAMIC COMPENSATION OF UREA AND TRIMETHYLAMINE N-OXIDE INTERACTIONS WITH PROTEIN
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DOI：
10.1021/bi00208a021
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发表时间：
1994-10-25
期刊：
影响因子：
2.9
通讯作者：
TIMASHEFF, SN
中科院分区：
文献类型：
--
作者：
LIN, TY;TIMASHEFF, SN
许多生物体在经历环境水胁迫时会积累称为渗透剂的低分子量物质。渗透剂的主要类别是糖、多元醇、氨基酸及其衍生物以及甲胺，并且所有这些都已知是蛋白质稳定剂。然而，海洋软骨鱼和腔棘鱼使用尿素和甲胺的组合作为渗透剂，即摩尔比为 2:1 的变性剂和稳定剂。使用核糖核酸酶 T1 (RNase T1) 作为蛋白质，在每种共溶剂单独存在及其混合物中进行优先结合和热变性测量。当尿素和 N-氧化三甲胺 (TMAO) 的摩尔比为 2:1 时，两种共溶剂对转变温度 (T-m) 的影响基本上是它们单独使用时影响的代数和。对尿素、TMAO 以及尿素与 TMAO 摩尔比为 2:1 的混合物的优先相互作用测量表明，TMAO 的存在对尿素与天然或未折叠（还原的羧甲基化 RNase T1）状态的蛋白质的相互作用没有影响。由于技术原因，无法测量 TMAO 在尿素存在下的优先相互作用。变性反应两个终态转移自由能的计算表明，无论是否存在 1 M TMAO，2 M 尿素都会使 RNase T1 不稳定 3.8 +/- 0.3 kcal/mol。在 2 M 尿素存在的情况下，1 M TMAO 对稳定性的贡献计算为 3.1 kcal/mol，在不存在 2 M 尿素的情况下测量为 2.7 kcal/mol。
Many organisms accumulate low molecular weight substances known as osmolytes when they experience environmental water stress. The main classes of osmolytes are sugars, polyhydric alcohols, amino acids and their derivatives, and methylamines, and all are known to be protein stabilizers. However, marine cartilaginous fishes and the coelacanth use, as osmolytes, a combination of urea and methylamines, i.e., a denaturant and a stabilizer, in a 2:1 molar ratio. Preferential binding and thermal denaturation measurements in the presence of each cosolvent separately and in their mixtures have been carried out using ribonuclease T1 (RNase T1) as the protein. At a 2:1 molar ratio of urea and trimethylamine N-oxide (TMAO), the effects of the two cosolvents on the transition temperature (T-m) were found to be essentially the algebraic sum of their effects when used individually. Preferential interaction measurements of urea, TMAO and urea in its 2:1 molar ratio mixture with TMAO, have shown that the presence of TMAO has no effect on the interaction of urea with the protein in either the native or the unfolded (reduced carboxymethylated RNase T1) state. The preferential interaction of TMAO in the presence of urea could not be measured for technical reasons. Calculations of transfer free energy in the two end states of the denaturation reaction have shown that 2 M urea destabilizes RNase T1 by 3.8 +/- 0.3 kcal/mol whether 1 M TMAO is present or not. The contribution of 1 M TMAO to stabilization is calculated to be 3.1 kcal/mol in the presence of 2 M urea and is measured to be 2.7 kcal/mol in its absence.