8TG8

Structure of Red beta C-terminal domain in complex with SSB C-terminal peptide, Form 3

8TG8 の概要

• エントリーDOI: 10.2210/pdb8tg8/pdb
• 分子名称: Recombination protein bet, TRP-MET-ASP-PHE-ASP-ASP-ASP-ILE-PRO-PHE (3 entities in total)
• 機能のキーワード: recombination, recombineering, single strand annealing, single-stranded dna binding protein, genome engineering, dna binding protein
• 由来する生物種: Escherichia phage Lambda; 詳細
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 10729.02

構造登録者

Bell, C.E. (登録日: 2023-07-12, 公開日: 2024-03-13, 最終更新日: 2024-05-15)

主引用文献

Zakharova, K.,Liu, M.,Greenwald, J.R.,Caldwell, B.C.,Qi, Z.,Wysocki, V.H.,Bell, C.E.
Structural Basis for the Interaction of Red beta Single-Strand Annealing Protein with Escherichia coli Single-Stranded DNA-Binding Protein.
J.Mol.Biol., 436:168590-168590, 2024

PubMed Abstract: Redβ is a protein from bacteriophage λ that binds to single-stranded DNA (ssDNA) to promote the annealing of complementary strands. Together with λ-exonuclease (λ-exo), Redβ is part of a two-component DNA recombination system involved in multiple aspects of genome maintenance. The proteins have been exploited in powerful methods for bacterial genome engineering in which Redβ can anneal an electroporated oligonucleotide to a complementary target site at the lagging strand of a replication fork. Successful annealing in vivo requires the interaction of Redβ with E. coli single-stranded DNA-binding protein (SSB), which coats the ssDNA at the lagging strand to coordinate access of numerous replication proteins. Previous mutational analysis revealed that the interaction between Redβ and SSB involves the C-terminal domain (CTD) of Redβ and the C-terminal tail of SSB (SSB-Ct), the site for binding of numerous host proteins. Here, we have determined the x-ray crystal structure of Redβ CTD in complex with a peptide corresponding to the last nine residues of SSB (MDFDDDIPF). Formation of the complex is predominantly mediated by hydrophobic interactions between two phenylalanine side chains of SSB (Phe-171 and Phe-177) and an apolar groove on the CTD, combined with electrostatic interactions between the C-terminal carboxylate of SSB and Lys-214 of the CTD. Mutation of any of these residues to alanine significantly disrupts the interaction of full-length Redβ and SSB proteins. Structural knowledge of this interaction will help to expand the utility of Redβ-mediated recombination to a wider range of bacterial hosts for applications in synthetic biology.

PubMed: 38663547
DOI: 10.1016/j.jmb.2024.168590

実験手法

X-RAY DIFFRACTION (1.576 Å)