Effect of dextran on protein stability and conformation attributed to macromolecular crowding
Effect of dextran on protein stability and conformation attributed to macromolecular crowding
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DOI：
10.1016/s0022-2836(02)01443-2
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发表时间：
2003-02-28
影响因子：
5.6
通讯作者：
Minton, AP
中科院分区：
文献类型：
--
作者：
Sasahara, K;McPhie, P;Minton, AP
用近紫外和远紫外圆二色谱法测定了鸡蛋清溶菌酶在不同浓度葡聚糖存在下的热诱导转变曲线。通过非线性最小二乘法将转变曲线拟合到两态模型，以获得展开转变的转变温度（T-m）、焓变（Δ H（u）（T-m））和自由能变（Δ G（u）（T））。在浓度超过约100 g l（-1）的葡聚糖存在下，观察到T-m增加和几乎恒定的Δ H（u）（T-m）值。此外，通过圆二色谱研究了葡聚糖诱导的完全未折叠蛋白质构象变化。在pH 2.0的酸未折叠的细胞色素c的溶液中添加高浓度的葡聚糖导致CD光谱从完全未折叠的多肽的特征转变为更紧凑的盐诱导的熔融球状态的特征，结果表明熔融球样状态相对于在拥挤的环境中完全未折叠的形式是稳定的。这两个观察是在定性雅阁与先前提出的模型的预测的影响，分子间排除体积（大分子拥挤）蛋白质的稳定性和构象。(C)2003爱思唯尔科技有限公司版权所有。
Thermally induced transition curves of hen egg-white lysozyme were measured in the presence of several concentrations of dextran at pH 2.0 by near-UV and far-UV CD. The transition curves were fitted to a two-state model by a non-linear, least-squares method to obtain the transition temperature (T-m), enthalpy change (DeltaH(u)(T-m)), and free energy change (DeltaG(u)(T)) of the unfolding transition. An increase in T-m and almost constant DeltaH(u)(T-m) values were observed in the presence of added dextran at concentrations exceeding ca 100 g l(-1). In addition, dextran-induced conformational changes of fully unfolded protein were investigated by CD spectroscopy. Addition of high concentrations of dextran to solutions of acid-unfolded cytochrome c at pH 2.0 results in a shift of the CD spectrum from that characteristic of the fully unfolded polypeptide to that characteristic of the more compact, salt-induced molten globule state, a result suggesting that the molten globule-like state is stabilized relative to the fully unfolded form in crowded environments. Both observations are in qualitative accord with predictions of a previously proposed model for the effect of intermolecular excluded volume (macromolecular crowding) on protein stability and conformation. (C) 2003 Elsevier Science Ltd. All rights reserved.