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Crystal structure of the bacteriophage P2 integrase catalytic domain
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
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2015 (English)In: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 589, no 23, 3556-3563 p.Article in journal (Refereed) Published
Abstract [en]

Bacteriophage P2 is a temperate phage capable of integrating its DNA into the host genome by site-specific recombination upon lysogenization. Integration and excision of the phage genome requires P2 integrase, which performs recognition, cleavage and joining of DNA during these processes. This work presents the high-resolution crystal structure of the catalytic domain of P2 integrase, and analysis of several non-functional P2 integrase mutants. The DNA binding area is characterized by a large positively charged patch, harbouring key residues. The structure reveals potential for large dimer flexibility, likely essential for rearrangement of DNA strands upon integration and excision.

Place, publisher, year, edition, pages
2015. Vol. 589, no 23, 3556-3563 p.
Keyword [en]
Bacteriophage P2, Integrase, Integration, Site-specific recombination, Tyrosine recombinase
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
URN: urn:nbn:se:su:diva-119219DOI: 10.1016/j.febslet.2015.09.026ISI: 000367232100008OAI: oai:DiVA.org:su-119219DiVA: diva2:844003
Available from: 2015-08-03 Created: 2015-08-03 Last updated: 2017-12-04Bibliographically approved
In thesis
1. Structural and biochemical studies of phage P2 DNA-binding proteins and human tetraspanins
Open this publication in new window or tab >>Structural and biochemical studies of phage P2 DNA-binding proteins and human tetraspanins
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Biochemical studies of proteins are crucial for a more detailed view of the world around us. The focus of biochemical studies can vary, from a complex mammalian system to a more simple viral entity, but the same methods and principles apply. In biochemistry one rely on both in vitro and in vivo analyses to understand biological processes. Protein crystallography has since the late 1950s been an additional important tool. By visualizing the structures of molecules involved in a biological process one can truly comprehend the molecular mechanisms of an organism or cell at the chemical level. This thesis includes structural biochemical work in combination with mutational and functional studies of proteins from both human and virus.

Human tetraspanins are integral membrane proteins grouped by their conserved structural features. Many of them have been shown to regulate cell migration, fusion, and signalling in the cell by functioning as organizers of multi-molecular membrane complexes. Several tetraspanins are also implicated in different forms of human cancers. How tetraspanins perform their function is still not known at the molecular level and today very little structural data exist on complete tetraspanin proteins. Structural biochemical studies require mg quantities of purified protein, something that is not easily obtained for membrane proteins. This thesis includes a family-wide approach to achieve full-length tetraspanins for biochemical studies. To facilitate this process a GFP-based optimization scheme for production and purification of membrane proteins in E. coli and S. cerevisiae has been applied. By utilizing this approach, we identified 8 human tetraspanins that can be produced and isolated from either E. coli or S. cerevisiae, and in one case using either system.

The temperate bacteriophage P2 is a virus, which can enter both the lytic and the lysogenic cycle upon infection of its host. The outcome of the infection is regulated by and dependent on several proteins encoded by the viral genome. The immunity repressor P2 and the Cox repressor direct the phage into either cycle. Integration and excision of the virus DNA requires the enzyme P2 integrase. The work in this thesis presents high-resolution crystal structures of these key proteins from the regulation of lysogeny in bacteriophage P2. By using a crystallographic approach in combination with mutational studies, key characteristics of these three proteins are presented. 

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2015. 52 p.
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-119225 (URN)978-91-7649-209-3 (ISBN)
Public defence
2015-09-18, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 3: Manuscript.

Available from: 2015-08-27 Created: 2015-08-03 Last updated: 2015-08-20Bibliographically approved

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Skaar, KarinClaesson, MagnusOdegrip, RichardHögbom, MagnusHaggård-Ljungquist, ElisabethStenmark, Pål
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Department of Biochemistry and BiophysicsDepartment of Molecular Biosciences, The Wenner-Gren Institute
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