Department of Life Sciences and Institute of Genome Sciences, National Yang-Ming University , 112 Taiwan.
Biochemistry. 2013 Nov 5;52(44):7714-22. doi: 10.1021/bi4010478. Epub 2013 Oct 23.
Thermophilic and hyperthermophilic archaea have one or more copies of the Alba gene, which encodes Alba, a dimeric, highly basic protein that binds cooperatively to DNA. However, the functions of Alba and how it interacts with DNA remain unclear. In this study, we have used single-molecule tethered particle motion (TPM) and optical tweezers (OT) experiments to study the interactions between DNA molecules and Alba1. When Alba1 binds to double-stranded DNA, the Brownian motion (BM) amplitude for DNA tethers increases continuously, suggesting that Alba1 binds cooperatively. The OT study confirmed that a 5-fold increase in the persistence length of the Alba1 nucleoprotein filament is the major factor causing the increase in the BM amplitude for DNA tethers, while the contour length remained mostly unchanged. Moreover, the rate of the increase in the BM amplitude and the BM plateau value are both DNA length-dependent, indicating that the number of Alba1 initiation binding sites increases as the DNA becomes longer. Using the incoming-strand TPM experiment to monitor the interaction between Alba1 nucleoprotein filaments, we found that significant dimer-dimer contacts between two Alba1 nucleoprotein filaments are present, and the interaction is regulated by the concentration of Alba1.
嗜热和超嗜热古菌具有一个或多个 Alba 基因的拷贝，该基因编码 Alba，一种二聚体、高度碱性的蛋白质，可协同结合 DNA。然而，Alba 的功能及其与 DNA 的相互作用仍不清楚。在这项研究中，我们使用单分子系绳粒子运动（TPM）和光学镊子（OT）实验来研究 DNA 分子与 Alba1 之间的相互作用。当 Alba1 结合双链 DNA 时，DNA 系绳的布朗运动（BM）幅度不断增加，表明 Alba1 结合具有协同性。OT 研究证实，Alba1 核蛋白丝的持久长度增加 5 倍是导致 DNA 系绳 BM 幅度增加的主要因素，而轮廓长度基本保持不变。此外，BM 幅度增加的速率和 BM 平台值均与 DNA 长度有关，表明随着 DNA 变长，Alba1 的起始结合位点数量增加。使用进入链 TPM 实验来监测 Alba1 核蛋白丝之间的相互作用，我们发现两个 Alba1 核蛋白丝之间存在明显的二聚体-二聚体接触，并且该相互作用受 Alba1 浓度的调节。
