荧光单链DNA结合蛋白可用于体外和体内研究。
Fluorescent single-stranded DNA-binding proteins enable in vitro and in vivo studies.
作者信息
Bianco Piero R, Stanenas Adam J, Liu Juan, Cohan Christopher S
机构信息
Center for Single Molecule Biophysics, Department of Microbiology and Immunology, University at Buffalo, Buffalo, NY, USA.
出版信息
Methods Mol Biol. 2012;922:235-44. doi: 10.1007/978-1-62703-032-8_18.
Abstract
Fluorescent single-stranded DNA-binding proteins (SSB) that have a defined number of fluorophores per tetramer are invaluable tools to understand biochemical mechanism and biological function. Here, we describe the purification of fluorescent SSB chimeras with a unique number of fluorescent subunits incorporated per tetramer. We describe the use of these tetramers to enable clear visualization of SSB in vivo. Purified chimeras also facilitate single molecule studies (Liu et al., Protein Sci 20:1005-1020, 2011).
摘要
每个四聚体含有特定数量荧光团的荧光单链DNA结合蛋白（SSB）是理解生化机制和生物学功能的宝贵工具。在此，我们描述了荧光SSB嵌合体的纯化方法，该嵌合体每个四聚体包含独特数量的荧光亚基。我们还描述了如何使用这些四聚体在体内清晰地可视化SSB。纯化的嵌合体也有助于单分子研究（Liu等人，《蛋白质科学》20:1005 - 1020，2011）。
相似文献
1
Fluorescent single-stranded DNA-binding proteins enable in vitro and in vivo studies.荧光单链DNA结合蛋白可用于体外和体内研究。
Methods Mol Biol. 2012;922:235-44. doi: 10.1007/978-1-62703-032-8_18.
2
Novel, fluorescent, SSB protein chimeras with broad utility.新型荧光 SSB 蛋白嵌合体具有广泛的应用。
Protein Sci. 2011 Jun;20(6):1005-20. doi: 10.1002/pro.633. Epub 2011 Apr 27.
3
Fluorescent SSB as a reagentless biosensor for single-stranded DNA.
Methods Mol Biol. 2012;922:219-33. doi: 10.1007/978-1-62703-032-8_17.
4
Mechanism of interaction between single-stranded DNA binding protein and DNA.单链 DNA 结合蛋白与 DNA 相互作用的机制。
Biochemistry. 2010 Feb 9;49(5):843-52. doi: 10.1021/bi901743k.
5
Application of the SSB biosensor to study in vitro transcription.SSB生物传感器在体外转录研究中的应用。
Biochem Biophys Res Commun. 2018 Feb 12;496(3):820-825. doi: 10.1016/j.bbrc.2018.01.147. Epub 2018 Jan 31.
6
A mutation in E. coli SSB protein (W54S) alters intra-tetramer negative cooperativity and inter-tetramer positive cooperativity for single-stranded DNA binding.大肠杆菌单链结合蛋白（W54S）中的一个突变改变了四聚体内的负协同性以及四聚体间单链DNA结合的正协同性。
Biophys Chem. 1997 Feb 28;64(1-3):235-51. doi: 10.1016/s0301-4622(96)02223-5.
7
Kinetic mechanism of direct transfer of Escherichia coli SSB tetramers between single-stranded DNA molecules.大肠杆菌单链结合蛋白（SSB）四聚体在单链DNA分子间直接转移的动力学机制
Biochemistry. 2002 Oct 1;41(39):11611-27. doi: 10.1021/bi020361m.
8
Single-molecule analysis of SSB dynamics on single-stranded DNA.单链DNA上SSB动力学的单分子分析
Methods Mol Biol. 2012;922:85-100. doi: 10.1007/978-1-62703-032-8_5.
9
SSB-DNA binding monitored by fluorescence intensity and anisotropy.通过荧光强度和各向异性监测单链结合蛋白与DNA的结合
Methods Mol Biol. 2012;922:55-83. doi: 10.1007/978-1-62703-032-8_4.
10
Use of fluorescently tagged SSB proteins in in vivo localization experiments.荧光标记的单链结合蛋白在体内定位实验中的应用。
Methods Mol Biol. 2012;922:245-53. doi: 10.1007/978-1-62703-032-8_19.
引用本文的文献
1
Selective engineering of condensation properties of single-stranded DNA binding (SSB) protein via its intrinsically disordered linker region.通过其内在无序的连接区对单链DNA结合（SSB）蛋白的凝聚特性进行选择性工程改造。
Nucleic Acids Res. 2025 Jun 6;53(11). doi: 10.1093/nar/gkaf481.
2
In Vivo Binding of Single-Stranded DNA-Binding Protein to Stalled Replication Fork Helicases.单链DNA结合蛋白在体内与停滞复制叉解旋酶的结合
Methods Mol Biol. 2021;2281:81-91. doi: 10.1007/978-1-0716-1290-3_5.
3
Rad51 Nucleoprotein Filament Disassembly Captured Using Fluorescent Plasmodium falciparum SSB as a Reporter for Single-Stranded DNA.利用荧光疟原虫单链结合蛋白作为单链DNA报告分子捕获Rad51核蛋白丝的解聚
PLoS One. 2016 Jul 14;11(7):e0159242. doi: 10.1371/journal.pone.0159242. eCollection 2016.
本文引用的文献
1
Novel, fluorescent, SSB protein chimeras with broad utility.新型荧光 SSB 蛋白嵌合体具有广泛的应用。
Protein Sci. 2011 Jun;20(6):1005-20. doi: 10.1002/pro.633. Epub 2011 Apr 27.
2
SSB protein diffusion on single-stranded DNA stimulates RecA filament formation.单链DNA上的SSB蛋白扩散刺激RecA丝状体形成。
Nature. 2009 Oct 22;461(7267):1092-7. doi: 10.1038/nature08442. Epub 2009 Oct 11.
3
RecG interacts directly with SSB: implications for stalled replication fork regression.RecG 与单链结合蛋白（SSB）直接相互作用：对停滞复制叉倒退的影响。
Nucleic Acids Res. 2008 Dec;36(22):7029-42. doi: 10.1093/nar/gkn795. Epub 2008 Nov 5.
4
SSB as an organizer/mobilizer of genome maintenance complexes.单链结合蛋白作为基因组维持复合物的组织者/动员者。
Crit Rev Biochem Mol Biol. 2008 Sep-Oct;43(5):289-318. doi: 10.1080/10409230802341296.
5
Fluorescent single-stranded DNA binding protein as a probe for sensitive, real-time assays of helicase activity.荧光单链DNA结合蛋白作为一种用于解旋酶活性灵敏实时检测的探针。
Biophys J. 2008 Oct;95(7):3330-9. doi: 10.1529/biophysj.108.133512. Epub 2008 Jul 3.
6
Resolution of converging replication forks by RecQ and topoisomerase III.RecQ和拓扑异构酶III对收敛复制叉的解析
Mol Cell. 2008 Jun 20;30(6):779-89. doi: 10.1016/j.molcel.2008.04.020.
7
Independent positioning and action of Escherichia coli replisomes in live cells.大肠杆菌复制体在活细胞中的独立定位与作用
Cell. 2008 Apr 4;133(1):90-102. doi: 10.1016/j.cell.2008.01.044.
8
A central role for SSB in Escherichia coli RecQ DNA helicase function.单链结合蛋白（SSB）在大肠杆菌RecQ DNA解旋酶功能中起核心作用。
J Biol Chem. 2007 Jun 29;282(26):19247-58. doi: 10.1074/jbc.M608011200. Epub 2007 May 3.
9
Structure of the DNA binding domain of E. coli SSB bound to ssDNA.与单链DNA结合的大肠杆菌单链结合蛋白（SSB）的DNA结合结构域的结构。
Nat Struct Biol. 2000 Aug;7(8):648-52. doi: 10.1038/77943.
Zotero 插件