Marceau Aimee H
Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA.
Methods Mol Biol. 2012;922:1-21. doi: 10.1007/978-1-62703-032-8_1.
Double-stranded (ds) DNA contains all of the necessary genetic information, although practical use of this information requires unwinding of the duplex DNA. DNA unwinding creates single-stranded (ss) DNA intermediates that serve as templates for myriad cellular functions. Exposure of ssDNA presents several problems to the cell. First, ssDNA is thermodynamically less stable than dsDNA, which leads to spontaneous formation of duplex secondary structures that impede genome maintenance processes. Second, relative to dsDNA, ssDNA is hypersensitive to chemical and nucleolytic attacks that can cause damage to the genome. Cells deal with these potential problems by encoding specialized ssDNA-binding proteins (SSBs) that bind to and stabilize ssDNA structures required for essential genomic processes. SSBs are essential proteins found in all domains of life. SSBs bind ssDNA with high affinity and in a sequence-independent manner and, in doing so, SSBs help to form the central nucleoprotein complex substrate for DNA replication, recombination, and repair processes. While SSBs are found in every organism, the proteins themselves share surprisingly little sequence similarity, subunit composition, and oligomerization states. All SSB proteins contain at least one DNA-binding oligonucleotide/oligosaccharide binding (OB) fold, which consists minimally of a five stranded beta-sheet arranged as a beta barrel capped by a single alpha helix. The OB fold is responsible for both ssDNA binding and oligomerization (for SSBs that operate as oligomers). The overall organization of OB folds varies between bacteria, eukaryotes, and archaea. As part of SSB/ssDNA cellular structures, SSBs play direct roles in the DNA replication, recombination, and repair. In many cases, SSBs have been found to form specific complexes with diverse genome maintenance proteins, often helping to recruit SSB/ssDNA-processing enzymes to the proper cellular sites of action. This clustering of genome maintenance factors can help to stimulate and coordinate the activities of individual enzymes and is also important for dislodging SSB from ssDNA. These features support a model in which DNA metabolic processes have evolved to work on ssDNA/SSB nucleoprotein filaments rather than on naked ssDNA. In this volume, methods are described to interrogate SSB-DNA and SSB-protein binding functions along with approaches that aim to understand the cellular functions of SSB. This introductory chapter offers a general overview of SSBs that focuses on their structures, DNA-binding mechanisms, and protein-binding partners.
双链（ds）DNA包含所有必要的遗传信息，尽管要实际利用这些信息需要解开双链DNA。DNA解旋会产生单链（ss）DNA中间体，这些中间体可作为众多细胞功能的模板。单链DNA的暴露给细胞带来了几个问题。首先，单链DNA在热力学上不如双链DNA稳定，这会导致双链二级结构的自发形成，从而阻碍基因组维护过程。其次，相对于双链DNA，单链DNA对化学和核酸酶攻击高度敏感，这些攻击可能会对基因组造成损害。细胞通过编码专门的单链DNA结合蛋白（SSB）来应对这些潜在问题，这些蛋白可结合并稳定基本基因组过程所需的单链DNA结构。SSB是在所有生命领域中都存在的必需蛋白。SSB以高亲和力且不依赖序列的方式结合单链DNA，通过这种方式，SSB有助于形成用于DNA复制、重组和修复过程的核心核蛋白复合底物。虽然在每种生物中都能找到SSB，但这些蛋白质本身在序列相似性、亚基组成和寡聚化状态方面却惊人地缺乏相似性。所有SSB蛋白都至少包含一个DNA结合寡核苷酸/寡糖结合（OB）折叠，它至少由一个五链β折叠组成，排列成一个由单个α螺旋封顶的β桶。OB折叠负责单链DNA结合和寡聚化（对于作为寡聚体发挥作用的SSB）。OB折叠的整体结构在细菌、真核生物和古细菌之间有所不同。作为SSB/单链DNA细胞结构的一部分，SSB在DNA复制、重组和修复中发挥直接作用。在许多情况下，已发现SSB与多种基因组维护蛋白形成特定复合物，通常有助于将SSB/单链DNA加工酶招募到适当的细胞作用位点。这种基因组维护因子的聚集有助于刺激和协调单个酶的活性，对于将SSB从单链DNA上解离也很重要。这些特征支持了一种模型，即DNA代谢过程已进化为作用于单链DNA/SSB核蛋白细丝，而不是裸单链DNA。在本卷中，描述了研究SSB - DNA和SSB - 蛋白质结合功能的方法以及旨在了解SSB细胞功能的方法。本章引言提供了对SSB的总体概述，重点关注其结构、DNA结合机制和蛋白质结合伙伴。
