Purified Stx and λ phage initiator O proteins bind specifically to two different origins of replication in vitro

Katarzyna I Kozłowska; Joanna Tymecka-Mulik; Grzegorz Węgrzyn

doi:10.1016/j.pep.2016.11.002

Purified Stx and λ phage initiator O proteins bind specifically to two different origins of replication in vitro

Protein Expr Purif. 2017 Mar:131:16-26. doi: 10.1016/j.pep.2016.11.002. Epub 2016 Nov 5.

Authors

Katarzyna I Kozłowska¹, Joanna Tymecka-Mulik², Grzegorz Węgrzyn²

Affiliations

¹ Department of Molecular Biology, University of Gdansk, Wita Stwosza 59, 80-308, Gdańsk, Poland. Electronic address: katarzyna.kozlowska@biol.ug.edu.pl.
² Department of Molecular Biology, University of Gdansk, Wita Stwosza 59, 80-308, Gdańsk, Poland.

PMID: 27826079
DOI: 10.1016/j.pep.2016.11.002

Abstract

The O protein is a crucial factor initiating the DNA replication of lambdoid bacteriophage. Efficient DNA replication of Shiga toxin-converting phage is necessary for effective production of Shiga toxin - main virulence factor of STEC strains. We developed an improved protocol for overproduction, bacterial cell lysis and purification of λO protein. With use of this method we have also isolated O proteins of Stx-phage P27 and 933W that were never purified before. Purified proteins were tested for their DNA binding activity and revealed a sequence specific interactions.

Keywords: DNA-Protein interactions; Initiator O protein; Protein overproduction; Protein purification; Stx-phage; λ phage.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Bacteriophage lambda / chemistry*
DNA, Viral / chemistry*
Protein Binding
Replication Origin*
Viral Proteins* / chemistry
Viral Proteins* / isolation & purification

Substances

DNA, Viral
O protein, Bacteriophage lambda
Viral Proteins