Journal article
Structure of a Holliday junction complex reveals mechanisms governing a highly regulated DNA transaction
eLife, v 5(2016)
25 May 2016
PMID: 27223329
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
The molecular machinery responsible for DNA expression, recombination, and compaction has been difficult to visualize as functionally complete entities due to their combinatorial and structural complexity. We report here the structure of the intact functional assembly responsible for regulating and executing a site-specific DNA recombination reaction. The assembly is a 240-bp Holliday junction (HJ) bound specifically by 11 protein subunits. This higher-order complex is a key intermediate in the tightly regulated pathway for the excision of bacteriophage λ viral DNA out of the E. coli host chromosome, an extensively studied paradigmatic model system for the regulated rearrangement of DNA. Our results provide a structural basis for pre-existing data describing the excisive and integrative recombination pathways, and they help explain their regulation.
Some viruses can remain dormant inside an infected cell and only become active when conditions are right to multiply and infect other cells. Bacteriophage λ is a much-studied model virus that adopts this lifecycle by inserting its genetic information into the chromosome of a bacterium called Escherichia coli. Certain signals can later trigger the viral DNA to be removed from the bacterial chromosome, often after many generations, so that it can replicate and make new copies of the virus. Specific sites on the viral and bacterial DNA earmark where the virus’s genetic information will insert and how it will be removed. Remarkably, each of these two site-specific reactions (i.e. insertion and removal) cannot be reversed once started, and their onset is precisely controlled.
These reactions involve a molecular machine or complex that consists of four enzymes that cut and reconnect the DNA strands and seven DNA-bending proteins that bring distant sites closer together. Despite decades of work by many laboratories, no one had provided a three-dimensional image of this complete molecular machine together with the DNA it acts upon.
Now, Laxmikanthan et al. reveal a three-dimensional structure of this machine with all its components by trapping and purifying the complex at the halfway point in the removal process, when the DNA forms a structure known as a “Holliday junction”. The structure was obtained using electron microscopy of complexes frozen in ice. The structure answers many of the long-standing questions about the removal and insertion reactions. For example, it shows how the DNA-bending proteins and enzymes assemble into a large complex to carry out the removal reaction, which is different from the complex that carries out the insertion reaction. It also shows that the removal and insertion reactions are each prevented from acting in the opposite direction because the two complexes have different requirements.
These new findings improve our understanding of how the insertion and removal reactions are precisely regulated. Laxmikanthan et al.’s results also serve as examples for thinking about the complicated regulatory machines that are widespread in biology.
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Details
- Title
- Structure of a Holliday junction complex reveals mechanisms governing a highly regulated DNA transaction
- Creators
- Gurunathan Laxmikanthan - Brown UniversityChen Xu - Brandeis UniversityAxel F Brilot - Brandeis UniversityDavid Warren - Brown UniversityLindsay Steele - Brown UniversityNicole Seah - Brown UniversityWenjun Tong - Brown UniversityNikolaus Grigorieff - Brandeis UniversityArthur Landy - Brown UniversityGregory D Van Duyne - University of Pennsylvania
- Publication Details
- eLife, v 5(2016)
- Publisher
- eLife
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- School of Biomedical Engineering, Science, and Health Systems
- Web of Science ID
- WOS:000387461400001
- Scopus ID
- 2-s2.0-84971668747
- Other Identifier
- 991019356339504721
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- Collaboration types
- Domestic collaboration
- Web of Science research areas
- Biology