Goobes Rivka, Kahana Nava, Cohen Orit, Minsky Abraham
Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel.
Biochemistry. 2003 Mar 4;42(8):2431-40. doi: 10.1021/bi026775x.
Crowding, which characterizes the interior of all living cells, has been shown to dramatically affect biochemical processes, leading to stabilization of compact morphologies, enhanced macromolecular associations, and altered reaction rates. Due to the crowding-mediated shift in binding equilibria toward association, crowding agents were proposed to act as a metabolic buffer, significantly extending the range of intracellular conditions under which interactions occur. Crowding may, however, impose a liability because, by greatly and generally enhancing macromolecular association, it can lead to irreversible interactions. To better understand the physical determinants and physiological consequences of crowding-mediated buffering, we studied the effects of crowding, or excluded volume, on DNA structures. Results obtained from isothermal titration calorimetry (ITC) and UV melting experiments indicate that crowding-induced effects are marginal under conditions that a priori favor association of DNA strands but become progressively larger when conditions deteriorate. As such, crowding exerts "genuine" buffering activity. Unexpectedly, crowding-mediated effects are found to include enthalpy terms that favorably contribute to association processes. We propose that these enthalpy terms and preferential stabilization derive from a reconfiguration of DNA hydration that occurs in dense DNA-rich phases obtained in crowded environments.
拥挤是所有活细胞内部的特征，已被证明会显著影响生化过程，导致紧密形态的稳定、大分子缔合增强以及反应速率改变。由于拥挤介导的结合平衡向缔合方向移动，拥挤剂被认为可作为一种代谢缓冲剂，显著扩展细胞内发生相互作用的条件范围。然而，拥挤可能带来不利影响，因为它通常会极大地增强大分子缔合，从而导致不可逆的相互作用。为了更好地理解拥挤介导的缓冲作用的物理决定因素和生理后果，我们研究了拥挤或排除体积对DNA结构的影响。等温滴定量热法（ITC）和紫外熔解实验结果表明，在先天有利于DNA链缔合的条件下，拥挤诱导的效应很小，但当条件变差时，这种效应会逐渐增大。因此，拥挤发挥着“真正的”缓冲活性。出乎意料的是，发现拥挤介导的效应包括对缔合过程有有利贡献的焓项。我们认为，这些焓项和优先稳定作用源于在拥挤环境中富含DNA的致密相中发生的DNA水合作用的重新配置。
