INFLUENCES OF MACROMOLECULAR CROWDING ON BIOCHEMICAL SYSTEMS

大肠杆菌DNA聚合酶I(POL I)的缺口翻译反应 被用作一个模型系统来演示 大分子拥挤以改变酶对 基本参数的数量，如pH、温度或抑制剂。 在存在高浓度的非特定聚合物的情况下， 尼克翻译发生在各种其他强烈的情况下 抑制条件。测试的条件包括一系列 PH值或温度或尿素的抑制浓度， 甲酰胺或溴化乙锭。这些拥挤效应是 在较高的离子强度下突出，表明它们起源于 增加聚合酶与其DNA模板之间的结合- 在拥挤的条件下使用底漆。动力学测量结果是 与这样的机制相一致。 我们研究了拥挤对协会的影响 核糖体颗粒的反应。约束的平衡点 大肠杆菌30个S核糖体亚基和50个S核糖体亚基的反应 在S存在下朝着形成70个核糖体的方向移动 各种聚合物。聚合物还进一步增加了 70个S粒子相互作用形成100个S二聚体。这个 对相对高浓度的非特定物质的要求 聚合物表明，平衡的变化是由 排除了音量效应。使用标度粒子理论的分析是 与这一机制一致。高浓度的影响 核糖体物种之间相互作用中的聚合物可能会使 拥挤环境下核糖体功能的重要变化 在体内发生的条件。

The nick-translation reaction of E. coli DNA polymerase I (Pol I) was used as a model system to demonstrate the ability of macromolecular crowding to alter the response of an enzyme to a number of basic parameters, such as pH, temperature or inhibitors. In the presence of high concentrations of non-specific polymers, nick translation occurred under a variety of otherwise strongly inhibitory conditions. The conditions tested included a range of pH values or temperatures or inhibitory concentrations of urea, formamide or ethidium bromide. These crowding effects are accentuated at higher ionic strengths, suggesting their origin in increased binding between the polymerase and its DNA template- primer under crowded conditions. Kinetic measurements were consistent with such a mechanism. We have examined the effects of crowding on the association reactions of ribosomal particles. The equilibrium for the binding reaction between the 30 S and 50 S ribosomal subunits of E. coli is shifted towards formation of 70 S ribosomes in the presence of a variety of polymers. The polymers also increase a further interaction between 70 S particles to form the 100 S dimer. The requirement for relatively high concentrations of non-specific polymers indicates that the shifts in equilibria arise from excluded volume effects. Analysis using scaled particle theory is consistent with this mechanism. The effects of high concentrations of polymers in the interactions between ribosomal species may make important changes in the function of ribosomes under the crowded conditions which occur in vivo.