皮牛秒力阶跃揭示了双链 DNA 延伸的转变动力学和机制。
PicoNewton-millisecond force steps reveal the transition kinetics and mechanism of the double-stranded DNA elongation.
机构信息
Laboratorio di Fisiologia, Dipartimento di Biologia Evoluzionistica, Università degli Studi di Firenze, Sesto Fiorentino, Italy.
出版信息
Biophys J. 2011 Aug 17;101(4):866-74. doi: 10.1016/j.bpj.2011.06.039.
Abstract
We study the kinetics of the overstretching transition in λ-phage double-stranded (ds) DNA from the basic conformation (B state) to the 1.7-times longer and partially unwound conformation (S state), using the dual-laser optical tweezers under force-clamp conditions at 25°C. The unprecedented resolution of our piezo servo-system, which can impose millisecond force steps of 0.5-2 pN, reveals the exponential character of the elongation kinetics and allows us to test the two-state nature of the B-S transition mechanism. By analyzing the load-dependence of the rate constant of the elongation, we find that the elementary elongation step is 5.85 nm, indicating a cooperativity of ~25 basepairs. This mechanism increases the free energy for the elementary reaction to ~94 k(B)T, accounting for the stability of the basic conformation of DNA, and explains why ds-DNA can remain in equilibrium as it overstretches.
摘要
我们使用双激光光学镊子在 25°C 下的力钳条件下，研究 λ 噬菌体双链 (ds) DNA 从基本构象 (B 态) 到拉长 1.7 倍且部分解旋的构象 (S 态) 的超伸展转变动力学。我们的压电力伺服系统具有前所未有的分辨率，可以施加 0.5-2 pN 的毫秒力阶，揭示了伸长动力学的指数特征，并使我们能够测试 B-S 转变机制的两态性质。通过分析伸长率的速率常数对负载的依赖性，我们发现基本伸长步骤为 5.85nm，表明协同性约为 25 个碱基对。该机制将基本反应的自由能增加到~94 k(B)T，解释了为什么 ds-DNA 在过度拉伸时仍能保持平衡。
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