Single-Molecule Analysis of Replication Protein A-DNA Interactions

被引：11

作者：

Bain, Fletcher E. ^{[1
]}

Fischer, Laura A. ^{[1
]}

Chen, Ran ^{[2
]}

Wold, Marc S. ^{[1
]}

机构：

[1] Univ Iowa, Carver Coll Med, Iowa City, IA 52242 USA

[2] Washington Univ, St Louis, MO USA

来源：

MECHANISMS OF DNA RECOMBINATION AND GENOME REARRANGEMENTS: METHODS TO STUDY HOMOLOGOUS RECOMBINATION | 2018年 / 600卷

关键词：

EXPRESSION; BINDING;

D O I：

10.1016/bs.mie.2017.11.016

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Replication protein A (RPA) is a highly conserved, eukaryotic ssDNA-binding protein essential for genome stability. RPA interacts with ssDNA and with protein partners to coordinate DNA replication, repair, and recombination. Single-molecule analysis of RPA-DNA interactions is leading to a better understanding of the molecular interactions and dynamics responsible for RPA function in cells. Here, we first describe how to express, purify, and label RPA. We then describe how to prepare materials and carry out single-molecule experiments examining RPA-DNA interactions using total internal reflection fluorescence microscopy (TIRFM). Finally, the last section describes how to analyze TIRFM data. This chapter will focus on human RPA. However, these methods can be applied to RPA homologs from other species.

引用

页码：439 / 461

页数：23

共 21 条

[1] Single-molecule sorting of DNA helicases [J].

Bain, Fletcher E. ;

Wu, Colin G. ;

Spies, Maria .

METHODS, 2016, 108 :14-23

[2] A minimal peptide substrate in biotin holoenzyme synthetase-catalyzed biotinylation [J].

Beckett, D ;

Kovaleva, E ;

Schatz, PJ .

PROTEIN SCIENCE, 1999, 8 (04) :921-929

[3] Functional assays for replication protein A (RPA) [J].

Binz, Sara K. ;

Dickson, Anne M. ;

Haring, Stuart J. ;

Wold, Marc S. .

DNA REPAIR, PT B, 2006, 409 :11-+

[4] Quantifying the Assembly of Multicomponent Molecular Machines by Single-Molecule Total Internal Reflection Fluorescence Microscopy [J].

Boehm, E. M. ;

Subramanyam, S. ;

Ghoneim, M. ;

Washington, M. Todd ;

Spies, M. .

SINGLE-MOLECULE ENZYMOLOGY: FLUORESCENCE-BASED AND HIGH-THROUGHPUT METHODS, 2016, 581 :105-145

[5]

BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3

[6] Introducing genetically encoded aldehydes into proteins [J].

Carrico, Isaac S. ;

Carlson, Brian L. ;

Bertozzi, Carolyn R. .

NATURE CHEMICAL BIOLOGY, 2007, 3 (06) :321-322

[7] Dynamic binding of replication protein a is required for DNA repair [J].

Chen, Ran ;

Subramanyam, Shyamal ;

Elcock, Adrian H. ;

Spies, Maria ;

Wold, Marc S. .

NUCLEIC ACIDS RESEARCH, 2016, 44 (12) :5758-5772

[8] Direct observation of individual RecA filaments assembling on single DNA molecules [J].

Galletto, Roberto ;

Amitani, Ichiro ;

Baskin, Ronald J. ;

Kowalczykowski, Stephen C. .

NATURE, 2006, 443 (7113) :875-878

[9] Direct Correlation of DNA Binding and Single Protein Domain Motion via Dual Illumination Fluorescence Microscopy [J].

Ghoneim, Mohamed ;

Spies, Maria .

NANO LETTERS, 2014, 14 (10) :5920-5931

[10] Cellular functions of human RPA1 - Multiple roles of domains in replication, repair, and checkpoints [J].

Haring, Stuart J. ;

Mason, Aaron C. ;

Binz, Sara K. ;

Wold, Marc S. .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2008, 283 (27) :19095-19111

← 1 2 3 →