Faithful cleavage of the P1 packaging site (pac) requires two phage proteins, PacA and PacB, and two Escherichia coli proteins, IHF and HU - PubMed (original) (raw)
. 1994 Oct 21;243(2):268-82.
doi: 10.1006/jmbi.1994.1653.
Affiliations
- PMID: 7932755
- DOI: 10.1006/jmbi.1994.1653
Faithful cleavage of the P1 packaging site (pac) requires two phage proteins, PacA and PacB, and two Escherichia coli proteins, IHF and HU
K Skorupski et al. J Mol Biol. 1994.
Abstract
The PacA and PacB subunits of the bacteriophage P1 DNA packaging enzyme (pacase) are necessary for cleavage of the phage packaging site (pac). In the accompanying paper, we show that the PacA subunit of the enzyme specifically binds to pac in the absence of PacB, but requires factors present in an Escherichia coli extract to do so. We show here that either of two E. coli DNA binding proteins, integration host factor (IHF) or HU, can replace this extract and promote the binding of PacA to pac. IHF binds to pac independently of PacA and DNase I footprinting experiments show that IHF protects approximately 40 bp of DNA around an IHF consensus sequence adjacent to the cleavage site. DNase I footprinting experiments also show that in the presence of either IHF or HU, PacA binds to the hexanucleotide sequences (5'-TGATCA/G) that flank the cleavage site and that have been previously shown to be essential for pac cleavage. The importance of IHF and HU in pac cleavage is further demonstrated by the severe reduction in both the fidelity and efficiency of pac cleavage in vitro with extracts lacking both IHF and HU. Addition of either IHF or HU to the deficient extracts renders them fully proficient for pac cleavage. Finally, we show that IHF bends DNA at the IHF site within pac. Based on these results, we propose a model that can account for the role of the various phage and host proteins, and for DNA bending in the pac cleavage reaction.
Similar articles
- Purification and DNA-binding activity of the PacA subunit of the bacteriophage P1 pacase enzyme.
Skorupski K, Sternberg N, Sauer B. Skorupski K, et al. J Mol Biol. 1994 Oct 21;243(2):258-67. doi: 10.1006/jmbi.1994.1652. J Mol Biol. 1994. PMID: 7932754 - IHF and HU: flexible architects of bent DNA.
Swinger KK, Rice PA. Swinger KK, et al. Curr Opin Struct Biol. 2004 Feb;14(1):28-35. doi: 10.1016/j.sbi.2003.12.003. Curr Opin Struct Biol. 2004. PMID: 15102446 Review. - Making DNA do a U-turn: IHF and related proteins.
Rice PA. Rice PA. Curr Opin Struct Biol. 1997 Feb;7(1):86-93. doi: 10.1016/s0959-440x(97)80011-5. Curr Opin Struct Biol. 1997. PMID: 9032059 Review.
Cited by
- P1 Bacteriophage-Enabled Delivery of CRISPR-Cas9 Antimicrobial Activity Against Shigella flexneri.
Huan YW, Torraca V, Brown R, Fa-Arun J, Miles SL, Oyarzún DA, Mostowy S, Wang B. Huan YW, et al. ACS Synth Biol. 2023 Mar 17;12(3):709-721. doi: 10.1021/acssynbio.2c00465. Epub 2023 Feb 20. ACS Synth Biol. 2023. PMID: 36802585 Free PMC article. - Bacteria have numerous distinctive groups of phage-plasmids with conserved phage and variable plasmid gene repertoires.
Pfeifer E, Moura de Sousa JA, Touchon M, Rocha EPC. Pfeifer E, et al. Nucleic Acids Res. 2021 Mar 18;49(5):2655-2673. doi: 10.1093/nar/gkab064. Nucleic Acids Res. 2021. PMID: 33590101 Free PMC article. - The genome sequence of Escherichia coli tailed phage D6 and the diversity of Enterobacteriales circular plasmid prophages.
Gilcrease EB, Casjens SR. Gilcrease EB, et al. Virology. 2018 Feb;515:203-214. doi: 10.1016/j.virol.2017.12.019. Epub 2018 Jan 2. Virology. 2018. PMID: 29304472 Free PMC article. - Structural and functional studies of the phage Sf6 terminase small subunit reveal a DNA-spooling device facilitated by structural plasticity.
Zhao H, Kamau YN, Christensen TE, Tang L. Zhao H, et al. J Mol Biol. 2012 Oct 26;423(3):413-26. doi: 10.1016/j.jmb.2012.07.016. Epub 2012 Jul 31. J Mol Biol. 2012. PMID: 22858866 Free PMC article. - DNA recombination with a heterospecific Cre homolog identified from comparison of the pac-c1 regions of P1-related phages.
Sauer B, McDermott J. Sauer B, et al. Nucleic Acids Res. 2004 Nov 18;32(20):6086-95. doi: 10.1093/nar/gkh941. Print 2004. Nucleic Acids Res. 2004. PMID: 15550568 Free PMC article.
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources